Synergistic compositions

ABSTRACT

The present invention describes a synergistic composition comprising of one or more statins, or one or more dipeptidyl peptidase IV (DPP IV) inhibitor or one or more biguanide antihyperglycaemic agent and a PPAR agonist of formula (Ia) 
     
       
         
         
             
             
         
       
     
     for the treatment of diabetes, especially non-insulin dependent diabetes (NIDDM) or Type 2 diabetes and conditions associated with diabetes mellitus and to compositions suitable for use in such method. The invention also describes the preparation of such compositions. The present invention also relates to certain novel salts of the PPAR agonist of formula (I): 
     
       
         
         
             
             
         
       
     
     processes for the preparation of these novel salts and use thereof.

FIELD OF THE INVENTION

The present invention describes a synergistic composition comprising ofone or more statins, or one or more dipeptidyl peptidase IV (DPP IV)inhibitor(s) or one or more biguanide antihyperglycaemic agent and aPPAR agonist of formula (I) for the treatment of diabetes, especiallynon-insulin dependent diabetes (NIDDM) or Type 2 diabetes and conditionsassociated with diabetes mellitus and to compositions suitable for usein such conditions. The invention also describes the preparation of suchcompositions. The present invention also relates to certain novel saltsof the PPAR agonist of formula (I), processes for the preparation ofthese novel salts and use thereof.

BACKGROUND OF THE INVENTION

Hyperlipidemia has been recognized as the major risk factor in causingcardiovascular diseases due to atherosclerosis. Atherosclerosis andother such peripheral vascular diseases affect the quality of life of alarge population in the world. The therapy aims to lower the elevatedplasma LDL cholesterol, low-density lipoprotein and plasma triglyceridesin order to prevent or reduce the risk of occurrence of cardiovasculardiseases.

Hypolipidemic agents which are PPAR modulators have been disclosed in WO91/19702, WO 94/01420, WO 94/13650, WO 95/03038, WO 95/17394, WO96/04260, WO 96/04261, WO 96/33998, WO 97/25042, WO 97/36579, WO98/28534, WO 99/08501, WO 99/16758, WO 99/19313, WO99/20614, WO00/23417, WO 00/23445, WO 00/23451, WO 01/53257.

WO 03009841 discloses compounds of the following general formula:

These compounds are reported to be hypolipidemic agents. WO 03009841also discloses certain salts including the sodium salt of some of thecompounds disclosed therein. However, these salts either were difficultto isolate due to rapid degradation or were poorly absorbed limitingtheir efficacy and possibility of further development or were found todegrade on long term storage thereby also limiting their suitability forfurther pharmaceutical development. It has surprisingly now been foundthat certain compounds and their selected salts are effective in furtherpharmaceutical development and are also efficacious than some of theearlier known salts.

The present invention provides synergistic composition of certainhypolipidemic and hypocholesterolemic compounds of formula (Ia) wherein‘M⁺’ represents Calcium, Magnesium, Sodium, Potassium, Zinc and Lithiumpreferably Magnesium in combination with one or more therapeutic agentsas described herein after.

In another embodiment the present invention describes compounds offormula (I)

wherein ‘R’ is selected from hydroxy, hydroxyalkyl, acyl, alkoxy,alkylthio, thioalkyl, aryloxy, arylthio and M⁺ represents Calcium,Magnesium, Sodium, Potassium, Zinc, Lithium, L-Arginine, Tromethamine,L-Lysine, Meglumine, Benethamine, Piperazine, Benzylamine,Dibenzylamine, Dicyclohexylamine, Diethylamine, Diphenylamine,α-naphthylamine, O-phenylenediamine, 1,3-Diaminopropane,(S)-α-naphthylethylamine, (S)-3-methoxyphenylethylamine,(S)-4-methoxyphenylethylamine, (S)-4-chlorophenylethylamine,(S)-4-methylphenylethylamine, Cinchonine, Cinchonidine, (−)-Quinine,Benzathine, Ethanolamine, Diethanol amine, Triethanolamine, imidazole,Diethylamine, Ethylenediamine, Choline, Epolamine, Morpholine4-(2-hydroxyethyl), N-N-diethylethanolamine, Deanol, Hydrabamine,Betaine, Ammonia, Adamantanamine, L-Adamantanmethylamine & Tritylamine.Many of these salts also shows certain superior pharmaceutical &/orchemical properties.

In a preferred embodiment, ‘R’ represents thioalkyl or alkoxy orhydroxyalkyl group; in a still preferred embodiment, ‘R’ represents—SCH₃ or —OCH₃ group.

Type 2 diabetes is a chronic and progressive disease arising from acomplex pathophysiology involving the dual endocrine defects of insulinresistance and impaired insulin secretion. The treatment of Type 2diabetes typically begins with diet and exercise, followed by oralantidiabetic monotherapy. For many patients, these regimens do notsufficiently control glycemia during long-term treatment, leading to arequirement for combination therapy within several years followingdiagnosis. However, co-prescription of two or more oral antidiabeticdrugs may result in treatment regimens that are complex and difficultfor many patients to follow. Combining two or more oral antidiabeticagents into a single tablet provides a potential means of deliveringcombination therapy without adding to the complexity of patients dailyregimens. Such formulations have been well accepted in other diseaseindications, such as hypertension (HYZAAR™ which is a combination oflosartan potassium and hydrochlorothiazide) and cholesterol lowering(VYTORIN™ which is a combination of simvastatin and ezetimibe). Theselection of effective and well-tolerated treatments is a key step inthe design of a combination tablet. Moreover, it is essential that thecomponents have complementary mechanisms of action and compatiblepharmacokinetic profiles. Examples of marketed combination tabletscontaining two oral antidiabetic agents include Glucovance™ (Metforminand Glyburide), Avandamet™ (Metformin and Rosiglitazone), Metaglip™(Metformin and Glipizide), Janumet™ (contains Sitagliptin and Metformin)etc.

Biguanide antihyperglycaemic agents are commonly used in the treatmentof NIDDM (or Type II diabetes). 1,1-Dimethylbiguanidine (or Metformin)is an example of a biguanides antihyperglycaemic agent.

Metformin represents the only oral antidiabetic agent proven to reducethe total burden of microvascular and macrovascular diabeticcomplications and to prolong the lives of Type 2 diabetic patients.Furthermore, metformin treatment is often associated with reductions inbody weight in overweight patients and with improvements in lipidprofiles in dyslipidemic patients. Metformin hydrochloride is marketedas either immediate-release or extended-release formulations with tabletdosage strengths of 500, 750, 850, and 1000 milligrams. Extended-releaseformulations of metformin have advantages over immediate-release interms of affording a more uniform maintenance of blood plasma activedrug concentrations and providing better patient compliance by reducingthe frequency of administration required.

It has now surprisingly been found that Compound of formula (Ia) incombination with a biguanide antihyperglycaemic agent such as Metforminprovides a particularly beneficial effect on glycaemic control with noobserved adverse effects; such combination is therefore particularlyuseful for the treatment of diabetes mellitus, especially Type IIdiabetes and conditions associated with diabetes mellitus.

Statins (or HMG-CoA reductase inhibitors) are a class of drugs used tolower cholesterol levels by inhibiting the enzyme HMG-CoA reductase,which plays a central role in the production of cholesterol in theliver. Increased cholesterol levels have been associated withcardiovascular diseases and statins are therefore used in the preventionof these diseases. Several statins have been approved for the treatmentof cardiovascular diseases including Lovastatin (MEVACOR, U.S. Pat. No.4,231,938), Simvastatin (ZOCOR; U.S. Pat. No. 4,444,784), Pravastatinsodium salt (PRAVACHOL; U.S. Pat. No. 4,346,227), Fluvastatin sodiumsalt (LESCOL; U.S. Pat. No. 5,354,772), Atorvastatin calcium salt(LIPITOR; U.S. Pat. No. 5,273,995) and Rosuvastatin calcium (CRESTOR;U.S. Pat. No. RE37314). The above mentioned publications areincorporated herein by reference.

The present invention is also based on the surprising finding thatstatins can increase the activity of PPAR agonists compound of formula(Ia) and can be used to treat or prevent dyslipidemia and type 2diabetes and other disorders responsive to PPAR activators or PPARactivation, without increasing the risk for side effects such asrhabdomylosis, fluid retention, edema, or congestive heart failure.

Dipeptidyl peptidase-4 (DPP-4) inhibitors represent a novel class ofagents that are being developed for the treatment or improvement inglycemic control in patients with Type 2 diabetes. Specific DPP-4inhibitors currently in clinical trials for the treatment of Type 2diabetes include sitagliptin phosphate (MK-0431), vildagliptin(LAF-237), saxagliptin (BMS-47718), alogliptin (X), carmegliptin (X),melogliptin (X), dutogliptin (X), denagliptin (X), linagliptin (X),P93/01 (Prosidion), SYR322 (Takeda), GSK 823093, Roche 0730699, TS021(Taisho), E3024 (Eisai), and PHX-1149 (Phenomix). For example, oraladministration of vildagliptin or sitagliptin to human Type 2 diabeticshas been found to reduce fasting glucose and postprandial glucoseexcursion in association with significantly reduced HbA_(1c) levels.Several reviews on the application of DPP-4 inhibitors for the treatmentof Type 2 diabetes, have been published such as H.-U. Demuth, et al.,“Type 2 diabetes—Therapy with dipeptidyl peptidase IV inhibitors,Biochim. Biophys. Acta, 1751: 33-44 (2005), K. Augustyns, et al.,“Inhibitors of proline-specific dipeptidyl peptidases: DPP IV inhibitorsas a novel approach for the treatment of Type 2 diabetes,” Expert Opin.Ther. Patents, 15: 1387-1407 (2005) etc.

DPP-4 inhibitors currently approved or are in clinical trials for thetreatment of Type 2 diabetes include sitagliptin phosphate,vildagliptin, saxagliptin, alogliptin, carmegliptin, melogliptin,dutogliptin, denagliptin, linagliptin, P93/01 (Prosidion), SYR322(Takeda), GSK 823093, Roche 0730699, TS021 (Taisho), E3024 (Eisai), andPHX-1149 (Phenomix) etc.

Sitagliptin free base and pharmaceutically acceptable salts thereof aredisclosed in U.S. Pat. No. 6,699,871 the contents of which are herebyincorporated by reference in their entirety. Crystalline sitagliptinphosphate monohydrate is disclosed in international patent publicationWO 2005/0031335.

Vildagliptin is the generic name for(S)-1-[(3-hydroxy-1-adamantyl)amino]acetyl-2-cyano-pyrrolidine.Vildagliptin is specifically disclosed in U.S. Pat. No. 6,166,063, thecontents of which are hereby incorporated by reference in theirentirety.

Saxagliptin is a methanoprolinenitrile derivative specifically disclosedin U.S. Pat. No. 6,395,767, the contents of which are herebyincorporated by reference in their entirety.

Alogliptin is a DPP-IV inhibitor under investigation for the treatmentof type 2 diabetes, specifically disclosed in EP 1586571 the contents ofwhich are hereby incorporated by reference in their entirety.

Linagliptin is a DPP-IV inhibitor approved for the treatment of type 2diabetes specifically disclosed in U.S. Pat. No. 7,407,955 the contentsof which are hereby incorporated by reference in their entirety.

Other DPP-IV inhibitors useful in the formulation of the presentinvention include, but are not limited to alogliptin, carmegliptin,melogliptin, dutogliptin, denagliptin.

The present invention is also based on the surprising finding that DPPIV inhibitors can increase the activity of PPAR agonists of formula (Ia)and can be used to treat or prevent dyslipidemia and type 2 diabetes andother disorders responsive to PPAR activators or PPAR activation,without increasing the risk for side effects such as rhabdomylosis,fluid retention, edema, or congestive heart failure.

Sodium-glucose co-transporter 2 (SGLT2) inhibitors are a new class ofdiabetic medications indicated only for the treatment of type 2diabetes. SGLT-2 inhibitors work by reducing the amount of glucose beingabsorbed in the kidneys so that it is passed out in the urine and alsoby reducing the amount of glucose in blood. Their use in clinicalpractice is associated with improved glycaemic control, weight loss anda low risk of hypoglycaemia. Dapagliflozin, Canagliflozin, Empagliflozinare some of the SGLT-2 inhibitors which are either approved or are underclinical trials for the treatment of diabetes and associated disorders.

The present invention is also based on the surprising finding thatSGLT-2 inhibitors can increase the activity of PPAR agonists compoundsof formula (Ia) and can be used to treat or prevent dyslipidemia andtype 2 diabetes and other disorders responsive to PPAR activators orPPAR activation, without increasing the risk for side effects such asrhabdomylosis, fluid retention, edema, or congestive heart failure.

Glucagone-like-peptide-1 agonists or GLP-1 agonists are a class of drugsfor the treatment of type 2 diabetes. Glucagon-like peptide-1 (GLP-1)enhances glucose-dependent insulin secretion following its release intothe circulation from the gut. GLP-1 receptor agonists enhanceglucose-dependent insulin secretion by the pancreatic beta-cell,suppress inappropriately elevated glucagon secretion, and slow gastricemptying. GLP-1 agonists are used for diabetes type 2 combined withother anti-diabetic drugs.

The present invention is also based on the surprising finding that GLP-1receptor agonists can increase the activity of PPAR agonists compoundsof formula (Ia) and can be used to treat or prevent dyslipidemia andtype 2 diabetes and other disorders responsive to PPAR activators orPPAR activation, without increasing the risk for side effects such asrhabdomylosis, fluid retention, edema, or congestive heart failure.

The acid of Formula (I) is a thick liquid which is difficult to isolate,purify and develop into a pharmaceutical formulation. It is thereforenecessary to isolate the acid in a form that is easy to purify, handle,scale up and develop into suitable pharmaceutical formulation.Conversion into suitable salts represent one such means.

Salts often improve physical and biological characteristics of mothercompounds without modifying primary pharmacological activity, based onmechanism of action of the compound. Thus there is a continuing need toobtain new salts of Formula (I) having improved physical and/or chemicalproperties. The present invention satisfies this need by providing newsalts of Formula (I).

In an embodiment, the new salts of Formula (I) provide a new opportunityto improve the performance of the synthesis of the Formula (I) acid in achemically and chirally pure form. These new salts are produced in solidstate, have improved characteristics such as stability, flowability andtherefore easy to handle in an industrial scale. This makes these newsalts suitable as intermediates for preparing the compound of formula(I) in a chemically and chirally pure form, though some of these saltsmay not be pharmaceutically useful. Some of these salts can also havesuperior biological properties over one or more of the known salts ofFormula (I).

These salts may be present either in substantially crystalline oramorphous forms or may be present as partially crystalline forms. In apreferred embodiment the salts are present in crystalline form. Inanother preferred embodiment, the salts are present in an amorphousform. In another embodiment, the salts are present innon-solvated/unsolvated form or in a solvent free form. In anotherembodiment, the salts are present in solvated/hydrated form.

SUMMARY OF THE PRESENT INVENTION

In one embodiment of the present invention is provided a synergisticcomposition comprising compound of formula (Ia) with one or more DPP IVinhibitors for the treatment of diabetes and its associated diseases.

In a further embodiment of the invention is provided a synergisticcomposition comprising compound of formula (Ia) with one or more statinsfor the treatment of diabetes and its associated diseases.

In a still further embodiment of the invention is provided a synergisticcomposition comprising compound of formula (Ia) with one or morebiguanides antihyperglycaemic agent for the treatment of diabetes andits associated diseases.

In another embodiment is provided a pharmaceutical compositioncomprising the compound of formula (Ia) and one or more therapeuticagents from those described above for the treatment of humans and othermammals in need thereof. In another embodiment is provided apharmaceutical composition comprising the compound of formula (Ia) andwith one or more thiazolidinedione antihyperglycaemic agents for thetreatment of humans and other mammals in need thereof.

In another embodiment is provided a pharmaceutical compositioncomprising the compound of formula (Ia) and with one or moresulfonylureas for the treatment of humans and other mammals in needthereof.

In another embodiment is provided a pharmaceutical compositioncomprising the compound of formula (Ia) and with one or more SGLT-2inhibitors for the treatment of humans and other mammals in needthereof.

In another embodiment is provided a pharmaceutical compositioncomprising the compound of formula (Ia) and with one or more insulinsensitizers for the treatment of humans and other mammals in needthereof.

In a still further embodiment is provided a pharmaceutical compositioncomprising, the therapeutically effective amount of compound of formula(Ia), prepared according to the present invention, along with at leastone suitable pharmaceutically acceptable carrier, diluents, vehicle orexcipient.

In another embodiment the present invention also provides novel salts ofcompound of Formula (I).

In another embodiment the present invention also provides processes forthe preparation of novel salts of compound of Formula (I).

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Calcium salt according to the present invention.

FIG. 2 is a powder X-ray diffraction pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Sodium salt according to the present invention.

FIG. 3 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Potassium salt according to the present invention.

FIG. 4 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Zinc salt according to the present invention.

FIG. 5 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Lithium salt according to the present invention.

FIG. 6 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Piperazine salt according to the present invention.

FIG. 7 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Tromethamine salt according to the present invention.

FIG. 8 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid L-Lysine salt according to the present invention.

FIG. 9 is a powder X-ray diffraction (XRPD) pattern of amorphous form of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Meglumine salt according to the present invention.

FIG. 10 is a powder X-ray diffraction (XRPD) pattern of crystalline formof (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Benethamine salt according to the present invention.

FIG. 11 is a powder X-ray diffraction (XRPD) pattern of crystalline formof (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Benzylamine salt according to the present invention.

FIG. 12 is a powder X-ray diffraction (XRPD) pattern of crystalline formof (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Dibenzylamine salt according to the present invention.

FIG. 13 is a powder X-ray diffraction (XRPD) pattern of amorphous formof (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Arginine salt according to the present invention.

FIG. 14 is a powder X-ray diffraction (XRPD) pattern of crystalline formof (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Imidazole salt according to the present invention.

DETAILED DESCRIPTION OF THE INVENTION

As used herein, the term “DIPE” refers to Diisopropyl ether; as usedherein the term “DMSO” refers to Dimethyl sulfoxide; as used herein, theterm “MTBE” refers to Methyl-tert Butyl Ether, as used herein, the term“THF” refers to Tetrahydrofuran; as used herein, the term “DMF” refersto N,N-Dimethyl formamide; as used herein, the term “DMA” refers toN,N-Dimethylacetamide; as used herein, the term “MIBK” refers to MethylIsobutyl Ketone; as used herein, the term “MEK” refers to Methyl ethylketone.

In one of the embodiment the present invention describes a synergisticcomposition comprising a compound of formula (Ia) and at least one moretherapeutic agent selected from one or more DPP IV inhibitors or one ormore biguanide antihyperglycaemic agents or one or more statins or oneor more thiazolidinediones or one or more sulfonylureas or one or moreSGLT2 inhibitors or one or more insulin sensitizers or one or more GLP-1agonists for the treatment of diabetes, dyslipidemia and theirassociated disorders.

wherein ‘M⁺’ represents Calcium, Magnesium, Sodium, Potassium, Zinc andLithium preferably Magnesium.

In another embodiment is provided compounds of Formula (I)

wherein ‘R’ is selected from hydroxy, hydroxyalkyl, acyl, alkoxy,alkylthio, thioalkyl, aryloxy, arylthio and M⁺ represents Calcium,Magnesium, Sodium, Potassium, Zinc, Lithium, L-Arginine, Tromethamine,L-Lysine, Meglumine, Benethamine, Piperazine, Benzylamine,Dibenzylamine, Dicyclohexylamine, Diethylamine, Diphenylamine,α-naphthylamine, O-phenylenediamine, 1,3-Diaminopropane,(S)-α-naphthylethylamine, (S)-3-methoxyphenylethylamine,(S)-4-methoxyphenylethylamine, (S)-4-chlorophenylethylamine,(S)-4-methylphenylethylamine, Cinchonine, Cinchonidine, (−)-Quinine,Benzathine, Ethanolamine, Diethanol amine, Triethanolamine, imidazole,Diethylamine, Ethylenediamine, Choline, Epolamine, Morpholine4-(2-hydroxyethyl), N-N-diethylethanolamine, Deanol, Hydrabamine,Betaine, Ammonia, Adamantanamine, L-Adamantanmethylamine, Tritylamine,Glucamine N-methyl, Pyrrolidine and the like.

In a preferred embodiment, ‘R’ represents thioalkyl, alkoxy orhydroxyalkyl group; in a still preferred embodiment, ‘R’ represents—SCH₃ or —OCH₃ group.

In a more preferred embodiment, the present invention discloses asynergistic composition comprising at least a second therapeutic agentselected from one or more DPP IV inhibitors or one or more biguanideantihyperglycaemic agents or one or more statins or one or morethiazolidinediones or one or more sulfonylureas or one or more SGLT2inhibitors and the compound of formula (Ia) wherein M⁺ representsMagnesium having the name Saroglitazar Magnesium & the chemical nameBenzenepropanoic acid,α-ethoxy-4-[2-[2-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]-,magnesium salt.

In an embodiment, the DPP IV inhibitors may be selected fromSitagliptin, Vildagliptin, Saxagliptin, Alogliptin and Linagliptinincluding their pharmaceutically acceptable salts, polymorphs, solvatesand hydrates. Other DPP IV inhibitors which may be used includeCarmegliptin, Melogliptin, Dutogliptin, Denagliptin, P93/01 (Prosidion),SYR322 (Takeda), GSK 823093, Roche 0730699, TS021 (Taisho), E3024(Eisai), and PHX-1149 (Phenomix) and the like.

In a preferred embodiment, the DPP IV inhibitor is Sitagliptin.

In another preferred embodiment, the DPP IV inhibitor is Vildagliptin.

In another preferred embodiment, the DPP IV inhibitor is Saxagliptin.

In another preferred embodiment, the DPP IV inhibitor is Alogliptin.

In another preferred embodiment, the DPP IV inhibitor is Linagliptin.

In an embodiment, the statins may be selected from Lovastatin,Pravastatin, Fluvastatin, Simvastatin, Atorvastatin, Rosuvastatin andPitavastatin.

In a preferred embodiment, the statins is Lovastatin also known by theinnovator brand name Mevacor.

In another preferred embodiment, the statin is Pravastatin also known bythe brand name Pravachol.

In another preferred embodiment, the statin is Simvastatin also known bythe innovator brand name Zocor.

In another preferred embodiment, the statin is Atorvastatin also knownby the innovator brand name Lipitor.

In yet another preferred embodiment, the statin is Rosuvastatin alsoknown by the innovator brand name Crestor.

In yet another preferred embodiment, the statin is Fluvastatin alsoknown by the innovator brand name Lescol.

In a still further embodiment, the statin is Pitavastatin also known bythe innovator brand name Livalo.

In another embodiment, the suitable biguanide antihyperglycaemic agentis selected from Metformin, Buformin or Phenformin.

In a preferred embodiment, the biguanide antihyperglycaemic agent isMetformin.

In another embodiment, the SGLT-2 inhibitors are selected fromCanaglifiozin, Dapagliflozinor Empagliflozin, Ertugliflozin &Ipragliflozin.

In another embodiment, the GLP-1 receptor agonist is selected fromExenatide or Liraglutide & Dulaglutide.

Accordingly, the invention provides a method for the treatment ofdyslipidemia, hypertriglyceridemia and diabetes mellitus and conditionsassociated with these disorders in a mammal such as a human, whichmethod comprises administering an effective, non-toxic andpharmaceutically acceptable amount of a compound of formula (I) and atleast one second therapeutic agent selected from one or more DPP IVinhibitors, or a statin or a suitable biguanide antihyperglycaemicagent, to a mammal in need thereof.

The method comprises either co-administration of a compound of formula(I) and one or more DPP IV inhibitors, or a statin or a suitablebiguanide antihyperglycaemic agent or other therapeutic agent asdescribed in the specification, or the sequential administrationthereof.

Co-administration includes administration of a formulation whichincludes both a compound of formula (I) and one or more DPP IVinhibitors or a statin or a suitable biguanide antihyperglycaemic agent,or one or more thiazolidinediones or one or more sulfonylureas or one ormore SGLT2 inhibitors or one or more insulin sensitizers or theessentially simultaneous administration of separate formulations of eachagent.

In another aspect the invention provides the use of a compound offormula (I) and a second therapeutic agent selected from one or more DPPIV inhibitors, or a statin or a suitable biguanide antihyperglycaemicagent, or one or more SGLT2 inhibitors, or one or more GLP-1-receptoragonist along with another suitable therapeutic agent for use in themanufacture of a composition for the treatment of obesity, diabetesmellitus, especially Type 2 diabetes and conditions associated withdiabetes mellitus.

Suitably, the other therapeutic agent comprises one or more, generallyone or two, of an antidiabetic agent, an alpha glucosidase inhibitor, abiguanide, an insulin secretagogue or an insulin sensitiser orsulphonylureas, DPP IV inhibitor.

A further suitable antidiabetic agent is insulin.

A suitable alpha glucosidase inhibitor is acarbose.

Other suitable alpha glucosidase inhibitors are emiglitate and miglitol.A further suitable alpha glucosidase inhibitor is voglibose.

Suitable DPP IV inhibitors which can be used include Sitagliptin,Vildagliptin, Saxagliptin, Alogliptin, Carmegliptin, Melogliptin,Dutogliptin, Denagliptin, Linagliptin, P93/01 (Prosidion), SYR322(Takeda), GSK 823093, Roche 0730699, TS021 (Taisho), E3024 (Eisai), andPHX-1149 (Phenomix) and the like.

Suitable biguanides include metformin, buformin or phenformin,especially metformin.

Suitable insulin secretagogues include sulphonylureas.

Suitable sulphonylureas include glibenclamide, glipizide, gliclazide,glimepiride, tolazamide and tolbutamide. Further sulphonylureas includeacetohexamide, carbutamide, chlorpropamide, glibomuride, gliquidone,glisentide, glisolamide, glisoxepide, glyclopyamide and glycylamide.Also included is the sulphonylurea glipentide.

A further suitable insulin secretagogue is repaglinide. An additionalinsulin secretagogue is nateglinide.

Insulin sensitisers also include thiazolidinedione insulin sensitisers.

Suitable thiazolidinedione insulin sensitisers include(RS)-5-[4-(2-[methyl(pyridin-2-yl)amino]ethoxy)benzyl]thiazolidine-2,4-dione (orRosiglitazone),(+)-5-[[4-[(3,4-dihydro-6-hydroxy-2,5,7,8-tetramethyl-2H-1-benzopyran-2-yl)methoxy]phenyl]methyl]-2,4-thiazolidinedione (or troglitazone),5-[4-[(1-methylcyclohexyl)methoxy]benzyl]thiazolidine-2,4-dione (orciglitazone),5-[4-[2-(5-ethylpyridin-2-yl)ethoxy]benzyl]thiazolidine-2,4-dione (orpioglitazone) or5-[(2-benzyl-2,3-dihydrobenzopyran)-5-ylmethyl)thiazolidin-2,4-dione (orenglitazone)

The present invention is based on the surprising finding that either aDPP IV inhibitor or a statin or a biguanide antihyperglycaemic agent ora sulphonylurea or an SGLT-2 inhibitor increases the activity of thePPAR agonists of formula (I) and can be used to treat or preventdyslipidemia and type 2 diabetes and other disorders responsive to PPARactivators or PPAR activation, without increasing the risk for sideeffects such as rhabdomylosis, fluid retention, edema, or congestiveheart failure.

It will be understood that compound of formula (I) and one or more DPPIV inhibitors or a statin or a suitable biguanide antihyperglycaemicagent are each administered in a pharmaceutically acceptable form,including pharmaceutically acceptable derivatives such aspharmaceutically acceptable salts, esters and solvates thereof, asappropriate of the relevant pharmaceutically active agent. In certaininstances herein the names used for the DPP IV inhibitors or thebiguanide antihyperglycaemic agent or statins etc. may relate to aparticular pharmaceutical form of the relevant active agent. It will beunderstood that all pharmaceutically acceptable forms of the activeagents per se are encompassed by this invention. Suitablepharmaceutically acceptable forms of the DPP IV inhibitor or thebiguanide antihyperglycaemic agent or statins etc. depend upon theparticular agent being used but include known pharmaceuticallyacceptable forms of the particular agent chosen. Such derivatives arefound or are referred to in standard reference texts such as the Britishand US Pharmacopoeias, Remington's Pharmaceutical Sciences (MackPublishing Co.), Martindale The Extra Pharmacopoeia (London, ThePharmaceutical Press) (for example see the 31st Edition page 341 andpages cited therein) and the likes.

The compounds of formula (I) can be prepared by the general processesand examples disclosed in WO 03009841. The DPP IV inhibitors or thebiguanide antihyperglycaemic agent or statins and other therapeuticagents used in the invention may be prepared by processes known in theart including those publications referred to earlier in thespecification.

Certain of the compounds mentioned herein may contain one or more chiralcarbon atoms and hence can exist in two or more isomeric forms, all ofwhich are encompassed by the invention, either as individual isomers oras mixtures of isomers, including racemates. If any of the compoundsmentioned herein, in particular the compounds of (I), exist in one ofseveral tautomeric forms, all of them are encompassed by the inventionas individual tautomeric forms or as mixtures thereof.

When used herein the term ‘dyslipidemia’ include conditions associatedwith hypertriglyceridemia and/or hypercholesterolemia and/or high LDL-Cand/or low HDL-C and complications associated with them.

‘Conditions associated with dyslipidemia’ include atherosclerosis,hypertension and other disorders known to a person skilled in the art.

When used herein the term ‘conditions associated with diabetes’ includesthose conditions associated with diabetes mellitus itself andcomplications associated with diabetes mellitus.

‘Conditions associated with diabetes mellitus itself’ includehyperglycaemia, insulin resistance, including acquired insulinresistance and obesity. Further conditions associated with diabetesmellitus itself include hypertension and cardiovascular disease,especially atherosclerosis and conditions associated with insulinresistance. Conditions associated with insulin resistance includepolycystic ovarian syndrome and steroid induced insulin resistance andgestational diabetes. ‘Complications associated with dyslipidemia anddiabetes mellitus’ includes renal disease neuropathy and retinopathy.

‘Renal diseases’ include nephropathy, glomerulonephritis, glomerularsclerosis, nephrotic syndrome, hypertensive nephrosclerosis and endstage renal disease.

Diabetes mellitus is preferably Type 2 diabetes.

It is also considered that the treatment of the invention will effect animprovement, relative to the individual agents, of one or moreparameters which are therapeutically relevant including in the levels ofadvanced glycosylation end products (AGEs), and serum lipids includingtotal cholesterol, HDL-cholesterol, LDL-cholesterol includingimprovements in the ratios thereof in particular an improvement in serumlipids including total cholesterol, HDL-cholesterol, LDL-cholesterolincluding improvements in the ratios thereof.

In the treatment of the invention, the active medicaments are preferablyadministered in pharmaceutical composition form. As indicated above,such compositions can include both medicaments and one only one of themedicaments in combination with the compound of formula (I).

Accordingly, in one aspect the present invention also provides apharmaceutical composition comprising a compound of formula (I) and atleast a second therapeutic agent selected from one or more DPP IVinhibitors or one or more biguanide antihyperglycaemic agent or one ormore statins or one or more thiazolidinediones or one or moresulfonylureas or one or more SGLT2 inhibitors or one or more insulinsensitizers and pharmaceutically acceptable excipients.

Thus, in a further aspect, the invention also provides a process forpreparing a pharmaceutical composition comprising a compound of formula(I) and at least a second therapeutic agent selected from one or moreDPP IV inhibitors or one or more biguanide antihyperglycaemic agents orone or more statins or other therapeutic agents as described elsewherein the specification and pharmaceutically acceptable excipients, whichprocess comprises admixing the compound of formula (I) and a second oroptionally a third therapeutically active agent as described in thespecification with suitable pharmaceutically acceptable excipients.

The compositions are preferably in a unit dosage form in an amountappropriate for the relevant daily dosage.

As used herein the term ‘pharmaceutically acceptable’ use embraces bothhuman and veterinary use.

In the treatment the medicaments may be administered from 1 to 6 times aday, but most preferably 1 or 2 times per day.

Also, the dosages of each particular active agent in any givencomposition can as required vary within a range of doses known to berequired in respect of accepted dosage regimens for that compound.Dosages of each active agent can also be adapted as required to takeinto account advantageous effects of combining the agents as mentionedherein.

It will be understood that the compound of formula (I) and the DPP IVinhibitors, biguanide antihyperglycaemic agents an SGLT-2 inhibitor or asulphonylurea and statins are in a pharmaceutically acceptable: form,including pharmaceutically acceptable derivatives such aspharmaceutically acceptable salts, esters and solvates thereof, asappropriate to the relevant pharmaceutically active agent chosen.

The present invention also provides a pharmaceutical compositioncomprising a compound of formula (I) and one or more DPP IV inhibitorsor biguanide antihyperglycaemic agent or statins or one or morethiazolidinediones or one or more sulfonylureas or one or more SGLT2inhibitors or one or more insulin sensitizers and pharmaceuticallyacceptable excipients, for use as an active therapeutic substance.

In particular, the present invention provides a pharmaceuticalcomposition comprising a compound of formula (I) and at least a secondtherapeutic agent selected from one or more DPP IV inhibitors orbiguanide antihyperglycaemic agents or statins or one or morethiazolidinediones or one or more sulfonylureas or one or more SGLT2inhibitors or one or more insulin sensitizers and pharmaceuticallyacceptable excipients, for use in the treatment of dyslipidemia anddiabetes mellitus, especially Type 2 diabetes and conditions associatedwith them.

Usually the compositions are adapted for oral administration. However,they may be adapted for other modes of administration, for exampleparenteral administration, sublingual or transdermal administration.

The compositions may be in the form of tablets, capsules, powders,granules, lozenges, suppositories, reconstitutable powders, or liquidpreparations, such as oral or sterile parenteral solutions orsuspensions.

In order to obtain consistency of administration it is preferred that acomposition of the invention is in the form of a unit dose.

Unit dosage presentation forms for oral administration may be in tabletor capsule form and may as necessary contain conventional excipientssuch as binding agents, fillers, lubricants, glidants, disintegrants andwetting agents.

The solid oral compositions may be prepared by conventional methods ofblending, filling or tableting. Repeated blending operations may be usedto distribute the active agent throughout those compositions employinglarge quantities of fillers. Such operations are of course conventionalin the art. The tablets may be coated according to methods well known innormal pharmaceutical practice, in particular with an enteric coating.

Oral liquid preparations may be in the form of, for example, emulsions,syrups, or elixirs, or may be presented as a dry product forreconstitution with water or other suitable vehicle before use. Suchliquid preparations may contain conventional additives such assuspending agents, for example sorbitol, syrup, methyl cellulose,gelatin, hydroxyethylcellulose, carboxymethylcellulose, aluminiumstearate gel, hydrogenated edible fats; emulsifying agents, for examplelecithin, sorbitan monooleate, or acacia; non-aqueous vehicles, (whichmay include edible oils), for example almond oil, fractionated coconutoil, oily esters such as esters of glycerine, propylene glycol, or ethylalcohol; preservatives, for example methyl or propyl p-hydroxybenzoateor sorbic acid; and if desired conventional flavouring or colouringagents.

For parenteral administration, fluid unit dosage forms are preparedutilizing the compound and a sterile vehicle, and, depending on theconcentration used, can be either suspended or dissolved in the vehicle.In preparing solutions the compound can be dissolved in water forinjection and filter sterilized before filling into a suitable vial orampoule and sealing. Advantageously, adjuvants such as a localanesthetic, a preservative and buffering agent can be dissolved in thevehicle. To enhance the stability, the composition can be frozen afterfilling into the vial and the water removed under vacuum. Parenteralsuspensions are prepared in substantially the same manner, except thatthe active compound is suspended in the vehicle instead of beingdissolved, and sterilization cannot be accomplished by filtration. Thecompound can be sterilized by exposure to ethylene oxide beforesuspending in the sterile vehicle. Advantageously, a surfactant orwetting agent is included in the composition to facilitate uniformdistribution of the compound. Compositions may contain from 0.1% to 99%by weight, preferably from 10-60% by weight, of the active material,depending upon the method of administration. Examples of binding agentsinclude acacia, alginic acid, carboxymethylcellulose calcium,carboxymethylcellulose sodium, dextrates, dextrin, dextrose,ethylcellulose, gelatin, liquid glucose, guar gum, hydroxyethylcellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose,magnesium aluminium silicate, maltodextrin, methyl cellulose,polymethacrylates, polyvinylpyrrolidone, pregelatinised starch, sodiumalginate, sorbitol, starch, syrup, tragacanth.

Examples of fillers include calcium carbonate, calcium phosphate,calcium sulphate, carboxymethylcellulose calcium, carboxymethylcellulosesodium, compressible sugar, confectioner's sugar, dextrates, dextrin,dextrose, dibasic calcium phosphate dihydrate, dibasic calciumphosphate, fructose, glyceryl palmitostearate, glycine, hydrogenatedvegetable oil-type 1, kaolin, lactose, maize starch, magnesiumcarbonate, magnesium oxide, maltodextrin, mannitol, microcrystallinecellulose, polymethacrylates, potassium chloride, powdered cellulose,pregelatinised starch, sodium chloride, sorbitol, starch, sucrose, sugarspheres, talc, tribasic calcium phosphate, xylitol.

Examples of lubricants include calcium stearate, glyceryl monostearate,glyceryl palmitostearate, magnesium stearate, microcrystallinecellulose, sodium benzoate, sodium chloride, sodium lauryl sulphate,stearic acid, sodium stearyl fumarate, talc, zinc stearate.

Examples of glidants include colloidal silicon dioxide, powderedcellulose, magnesium trisilicate, silicon dioxide, talc.

Examples of disintegrants include alginic acid, carboxymethylcellulosecalcium, carboxymethylcellulose sodium, colloidal silicon dioxide,croscarmellose sodium, crospovidone, guar gum, magnesium aluminiumsilicate, microcrystalline cellulose, methyl cellulose,polyvinylpyrrolidone, polacrilin potassium, pregelatinised starch,sodium alginate, sodium lauryl sulphate, sodium starch glycollate.

An example of a pharmaceutically acceptable wetting agent is sodiumlauryl sulphate.

The compositions are prepared and formulated according to conventionalmethods, such as those disclosed in standard reference texts and arewell within the scope of a skilled person. For example, the solid oralcompositions may be prepared by conventional methods of blending,filling or tableting. Repeated blending operations may be used todistribute the active agent throughout those compositions employinglarge quantities of fillers. Such operations are of course conventionalin the art. The tablets may be coated according to methods well known innormal pharmaceutical practice.

Compositions may, if desired, be in the form of a pack accompanied bywritten or printed instructions for use.

No adverse toxicological effects were seen for the compositions ormethods of the invention in the above mentioned dosage ranges. Furtherthe composition of the present invention was found suitable for thetreatment of diabetes and its associated disorders without increasingthe risk for fluid retention, peripheral edema, pulmonary edema, orcongestive heart failure.

In one of the embodiment the present invention provides certain newsalts of compound of Formula (I)

wherein ‘R’ is selected from hydroxy, hydroxyalkyl, acyl, alkoxy,alkylthio, thioalkyl, aryloxy, arylthio and M⁺ represents Calcium,Sodium, Potassium, Zinc, Lithium, L-Arginine, Tromethamine, L-Lysine,Meglumine, Benethamine, Piperazine, Benzylamine, Dibenzylamine,Dicyclohexylamine, Diethylamine, Diphenylamine, α-naphthylamine,O-phenylenediamine, 1,3-Diaminopropane, (S)-α-naphthylethylamine,(S)-3-methoxyphenylethylamine, (S)-4-methoxyphenylethylamine,(S)-4-chlorophenylethylamine, (S)-4-methylphenylethylamine, Cinchonine,Cinchonidine, (−)-Quinine, Benzathine, Ethanolamine, Diethanol amine,Triethanolamine, imidazole, Diethylamine, Ethylenediamine, Choline,Epolamine, Morpholine 4-(2-hydroxyethyl), N-N-diethylethanolamine,Deanol, Hydrabamine, Betaine, Ammonia, Adamantanamine,L-Adamantanmethylamine, Tritylamine, Glucamine N-methyl, Pyrrolidine.

In a preferred embodiment, ‘R’ represents thioalkyl and alkoxy orhydroxyalkyl group; In a still preferred embodiment, ‘R’ represents—SCH₃ or —OCH₃ group.

In an embodiment these salts may be present either in crystalline oramorphous form or suitable mixtures of crystalline and amorphous forms.In a further embodiment, each of the crystalline and/or amorphous formsmay independently exist either in hydrated, solvated, non-solvated,anhydrous, solvent free or desolvated solvates of either thecrystalline, amorphous or various mixtures of crystalline and amorphousforms.

In one embodiment the some of the novel salts of the present inventioncan be used for the purification of free acid of formula (I) by reactingthe impure acid with suitable salt in a suitable solvent and then thepure acid is obtained from the salt by suitable techniques. The purefree acid of formula (I) can be further converted to Magnesium & othertherapeutic salts of compound of formula (I)

In one embodiment of the invention is provided the (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid calcium salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid calcium salt can be present in either crystalline and/or amorphousform each of which can optionally be present in anhydrous, solvent free,hydrated or solvated forms.

In a preferred embodiment, the calcium salt is present in crystallineform.

In another preferred embodiment, the calcium salt is present inamorphous form.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid sodium salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid sodium salt can be present in either crystalline and/or amorphousform each of which can optionally be present in anhydrous, hydrated orsolvated forms.

In a preferred embodiment, the sodium salt is present in crystallineform.

In another preferred embodiment, the sodium salt is present in amorphousform.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid potassium salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid potassium salt can be present in either crystalline and/oramorphous, form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In a preferred embodiment, the potassium salt is present in crystallineform.

In another preferred embodiment, the potassium salt is present inamorphous form.

In an embodiment is provided(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionic acid zinc salt. In anembodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid zinc salt can be present in either crystalline and/or amorphousform each of which can optionally be present in anhydrous, hydrated orsolvated forms.

In a preferred embodiment, the zinc salt is present in crystalline form.

In another preferred embodiment, the zinc salt is present in amorphousform.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid lithium salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid lithium salt can be present in either crystalline and/or amorphousform each of which can optionally be present in anhydrous, hydrated orsolvated forms.

In one embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid L-arginine salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid L-arginine salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Tromethamine salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Tromethamine salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In an embodiment is provided(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid L-Lysine salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid L-Lysine salt can be present in either crystalline and/or amorphousform each of which can optionally be present in anhydrous, hydrated orsolvated forms.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Meglumine salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Meglumine salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Benethamine salt. In a further embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Benethamine salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In a preferred embodiment is provided a crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Benethamine salt according to the present invention characterizedby its powder X-ray diffraction pattern having peaks expressed as 2θ±0.2degree at about 10.0, 10.3, 14.5, 15.1, 15.7, 16.7, 17.4, 17.9, 18.6,19.2, 19.8, 21.3, 23.2 and 25.7.

In a still further preferred embodiment the crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Benethamine salt is further characterized by its powder X-raydiffraction pattern (PXRD) as depicted in FIG. 10.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid piperazine salt. In an embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid piperazine salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionic acid benzylamine salt. Ina further embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid benzylamine salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In a preferred embodiment is provided a crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid benzylamine salt according to the present invention characterizedby its powder X-ray diffraction pattern having peaks expressed as 2θ±0.2degree at about 14.8, 16.8, 17.5, 18.3, 19.3, 20.8, 22.6 and 24.2.

In a still further preferred embodiment the crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid benzylamine salt is further characterized by its powder X-raydiffraction pattern (PXRD) as depicted in FIG. 11.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Dibenzylamine salt. In a further embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Dibenzylamine salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In a preferred embodiment is provided a crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Dibenzylamine salt according to the present invention characterizedby its powder X-ray diffraction pattern having peaks expressed as 2θ±0.2degree at about 8.72, 16.8, 18.5, 19.1, 19.6, 20.6, 21.6, 22.5 and 24.5.

In a still further preferred embodiment the crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Dibenzylamine salt is further characterized by its powder X-raydiffraction pattern (PXRD) as depicted in FIG. 12.

In an embodiment is provided (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid imidazole salt. In a further embodiment, (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid imidazole salt can be present in either crystalline and/oramorphous form each of which can optionally be present in anhydrous,hydrated or solvated forms.

In a preferred embodiment is provided a crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid imidazole salt according to the present invention characterized byits powder X-ray diffraction pattern having peaks expressed as 2θ±0.2degree at about 9.40, 14.7, 15.6, 17.3, 21.0, 21.5, 22.5 and 26.2.

In a still further preferred embodiment the crystalline form of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Imidazole salt is further characterized by its powder X-raydiffraction pattern (PXRD) as depicted in FIG. 14.

Analytical Methods:

i) Chromatographic Purity/Related Substance (by HLC)

Column: YMC-pack ODS-AM (type L₁) or equivalent; 250×4.6 mm, 5 μm

Wave length: 294 nm

Column temp.: 30° C.

Mobile phase: [Ammonium acetate buffer: Acetonitrile]/[55:45]

Flow rate: 1 ml/min

Injection vol.: 5 μl

Retention time: About 10 mints.

Run time: 60 mints.

ii) Chromatographic Purity Chiral (by HPLC)

Column: chiralcel OJ-H: 250×4.6 mm, 5 μm

Wave length: 294 nm

Column temp.: 35° C.

Mobile phase: 0.05% TFA in EtOH: Hexane/(12:88)

Flow rate: 0.8 ml/min

Injection vol.: 5 μl

Run time: 60 mints.

iii) DSC (Differential Scanning Calorimeter):

Weight accurately 2 mg to 3 mg of sample in a clean aluminum pan, placethe lid and seal it with the help of sealing process.

Instrument Detail:

Make: Perkin Elmer

Model: Pyris 1

Software: Pyris 1

Follow the method as describe below:

Initial temperature: 50° C.

Final temperature: 300° C.

Heating rate: 10° C./min

Gas: Nitrogen

Gas Flow: 20 ml/min

Blank Run:

Run the blank for baseline correction by placing empty aluminum pans inboth sample and reference compartments of the DSC furnace and run a scanusing the temperature range and at a heating rate mentioned inInstrumental parameters.

Sample Run:

Place the sample preparation in the sample compartment and blankaluminum pan in the reference compartment of the DSC furnace and run ascan using the temperature range and heating rate mentioned as ininstrumental parameters using the base line file obtained in the blankrun above for the baseline correction.

iv) XRPD Method

Sample Preparation:

Place a Sufficient quantity of sample to be analyzed on the sampleholder plate and flatten it with the help of another plate to achieve asmooth surface. Record the diffraction pattern as per below instrumentalparameters

Instrument used: 2 k W XRD

Model: MF2100

Make: Rigaku

Instrument Parameters:

1. X-ray: Cu/40 kV/30 mA

2. Diversion slit: 1°

3. Scattering slit: 1°

4. Receiving slit: 0.15 mm

5. Filter: Ni-kβ filter

6. Counter: Scintillation counter

7. Scan mode: Continuous

8. Scan speed: 4.000°/minute

9. Sampling width: 0.010°

10. Scan axis: 2theta\theta

11. Scan range: 2.00 to 40.0°

12. Theta offcet: 0.000°

v) NMR Analysis:

The 1H NMR spectrum & 13C NMR spectrum was obtained for Saroglitazarsalts in DMSO-d6 solvent by using Bruker AVANCE II 400 MHz spectrometerinstrument.

vi) Mass Spectrum Analysis:

The mass spectrum for Saroglitazar salts were obtained using LCMS 2010 ASHIMADZU instrument by positive/negative chemical ionization massspectrometry.

In one of the embodiments the salts of Formula (I) prepared according tothe present invention, preferably have a purity of at least 98%, morepreferably at least 99%.

In one embodiment, some of the salts of the present invention may beused as an intermediate to obtain highly pure acid in Formula (I)

In another embodiment, the salts of the present invention can beformulated to their suitable pharmaceutical compositions for use inhumans and other suitable species.

Accordingly, the invention provides a method for the treatment ofdyslipidemia, hypertriglyceridemia and diabetes mellitus and conditionsassociated with these disorders in a mammal such as a human, whichmethod comprises administering an effective, non-toxic andpharmaceutically acceptable amount of a compound of formula (I) and oneor more DPP IV inhibitors, to a mammal in need thereof.

In an embodiment the present invention provides an improved process forpreparing compound of formula (I) using novel intermediates.

The invention is further exemplified by the following non-limitingexamples, which are illustrative representing the preferred modes ofcarrying out the invention. The invention's scope is not limited tothese specific embodiments only but should be read in conjunction withwhat is disclosed anywhere else in the specification together with thoseinformation and knowledge which are within the general understanding ofa person skilled in the art.

Example-1 Preparation of (S)-α-1-phenylethylamine salt of(S)-α-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]benzenepropanoicacid

In a dry, 100 mL round bottom flask methanol (25 L) was taken andracemicα-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]benzenepropanoic acid ethyl ester (6.3 g) was added at room temperature, undernitrogen.

Sodium hydroxide (0.591 g) dissolved in water (6 mL) was added into thereaction mixture and stirred at room temperature for 3 hours to completehydrolysis. Solvent was removed under reduced pressure. Water (65 mL)was added to concentrate the material. Impurity was removed by solventwashing. Aqueous layer was acidified with acetic acid and extracted withisopropyl acetate. The layer of isopropyl acetate was dried over sodiumsulfate. Subsequently (S)-α-phenyl ethylamine (0.965 g) was added to thereaction. Solvent was distilled off to remove maximum isopropyl acetate.To this acetonitrile (55 mL) was added and stirred at room temperaturefor 4 hours. Solid filtered and dried.

Yield: 2.55 g, HPLC Purity: 97.17° %, Chiral purity: 94.19%.

Example-2 Preparation of (S)-α-1-phenylethylamine salt of(S)-α-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]benzenepropanoicacid

In a dry, 20 L round bottom flask methanol (6.868 L) was taken andracemicα-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]benzene-propanoicacid ethyl ester (1.717 g) was added at room temperature under nitrogen.Sodium hydroxide (153.95 g) dissolved in water (1.717 L) was added intothe reaction mixture and stirred at room temperature for 3 hours tocomplete hydrolysis. Solvent was removed under reduced pressure. Water(17.17 L) was added to concentrated material. Impurity was removed bysolvent washing. Aqueous layer was acidified with acetic acid andextracted with isopropyl acetate. Isopropyl acetate layer was dried oversodium sulfate. Subsequently (R)-α-phenyl ethylamine (287 g) was addedto the reaction. Solvent was distilled off to remove maximum isopropylacetate. To this, acetonitrile (27.22 L) was added and stirred at roomtemperature for 24 hours. Solid was filtered. To filtrate added(S)-α-phenyl ethylamine (269.07 g). Reaction mixture was stirred at roomtemperature for 4.5 hours. Solid was filtered and dried.

Yield: 603 g (72.76% yield), HPLC Purity: 99.21%, Chiral purity: 94.00%.

Example-3 Preparation of (S)-α-1-phenylethylamine salt of(S)-α-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]benzenepropanoicacid

In a dry, 20 L round bottom flask 3.640 L methanol was taken and 910 gracemicα-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]benzenepropanoic acid ethyl ester was added into the reaction mixture at roomtemperature under nitrogen. 84.38 g sodium hydroxide was dissolved in910 mL water and added into the reaction mixture and stirred at roomtemperature for 3 hours to complete hydrolysis. Solvent was removedunder reduced pressure. 9.1 L water was added to concentrate thematerial. Impurity was removed by solvent washing. Aqueous layer wasacidified with acetic acid and extracted with isopropyl acetate.Isopropyl acetate layer was dried over sodium sulfate. Subsequently196.65 g (R)-α-phenyl ethylamine was added to the reaction. Solvent wasdistilled off to remove maximum isopropyl acetate. To this 6.570 Lacetonitrile was added and stirred at room temperature for 24 hours.Solid was filtered. To the filtrate added 26.96 g (S)-1-(4-nitrophenyl)ethylamine and 176.98 g (S)-α-phenyl ethylamine. Reaction mixture wasstirred at room temperature for 4.5 hr. Solid was filtered and dried.

Yield: 338 g (74.38%), HPLC Purity: 98.16%, Chiral purity: 98.76%.

Similarly, (S)-α-1-phenylethylamine salt of(S)-α-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-yl]ethoxy]benzenepropanoicacid is prepared in different batches and the results are summarized intable 1 given below.

TABLE 1 Chemical Chiral No. Input Output % Yield purity purity 4  6.5 g 2.35 g 78.48% 99.43% 98.23% 5  6.5 g  2.3 g 76.82% 99.19% 97.94% 6 829g 355.7 g 97.56% 99.30% 92.61%

Purity (HPLC): 99.70%, Chiral purity: 99.57%

¹H NMR: 1.04 (3H, t, J=7.0 Hz, —CH₃), 1.49 (3H,d, J=6.8 Hz, —CH₃), 2.35(3H,s, —CH₃), 2.50 (3H,s, —SCH₃), 2.85 & 2.74 (2H,m, CH₂), 3.46 & 3.18(2H,m, CH₂), 3.76 (2H, m, —CH), 4.17 (111, q, —CH), 4.25 & 3.89 (4H, t,—CH₂ & t, —CH₂), 6.08 & 5.95 (d, J=3.27 Hz, 2×CH (pyrrole)), 6.58 (dd,J=8.8 Hz, 2×-ArH), 7.04 (dd, J=8.8 Hz, 2×ArH), 7.24 (m, 2×ArH), 7.26 (m,2×ArH), 7.28 (m, 2×ArH), 7.31 (m, 2×ArH), 7.38 (m, 2×ArH).

¹³C NMR: 12.95, 15.28, 15.89, 22.13, 38.77 43.28, 5120, 65.50, 66.97,82.31, 107.25, 108.54, 113.86, 126.65, 128.32, 128.94, 129.69, 130.29,130.42, 130.86, 132.09, 133.47, 137.22, 140.29, 156.62, 179.15.

ESIMS: 438.2 (M−1) (molecular ion peak of(S)-α-ethoxy-4-[2-[-methyl-5-[4-(methylthio)phenyl]-1H-pyrrol-1-yl]ethoxy]benzenepropanoicacid.

IR: Frequency (cm⁻¹): 3441.12, 2970.48, 2920.32, 1624

Example-7 Preparation of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid calcium salt

In 250 ml round bottom flask, (S,S)(−,−)-α-Methyl benzylamine (10.0 g,0.017 mole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy)-phenyl)-propionicacid was charged. A solution of ethyl acetate (72 ml) and water (50 ml)was added at RT under N₂ atm., 50% HCl (4.8 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out at rotavapor under vacuum at 50-55°C. Free acid compound was obtained. In a 250 ml three necked flask,above oily mass (free acid compd.) was charged and dissolved in methanol(39 ml), added a solution of Sodium hydroxide (0.929 g in 39 ml water)(0.023 mole) and stirred for 15 minutes at RT. The reaction mass wascooled at 10-15° C. and added a solution of calcium acetate (2.05 g in39 ml water) (0.011 mole) within 5 minutes. Salt was precipitated outand stirred it for 30 minutes. The solid was filtered and washed withwater. Dried the solid under vacuum. Yield: 8.0 g (97.8%), HPLC: purity:98.82%, Chiral purity: 99.52%.

M.P.: 175.70° C.

Example-8 Preparation of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid Sodium salt

In 250 ml round bottom flask, (S,S)(−,−)α-methyl benzylamine (10.0 g,0.017 mole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (72 ml) and water (50 ml)was added at RT under N₂ atm., 50% HCl (4.8 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum at50-55° C. Free acid compound was obtained. In a 250 ml three neckedflask, above oily mass (free acid compd.) charged and dissolved inMethanol (50 ml), added Sodium methoxide (0.868 g, 0.016 mole) undernitrogen atmosphere and stirred for 30 minutes at RT. The solvent wasdistilled out using rotavapor. Solid material was obtained. Trituratedthe solid material with n-Heptane and stirred for 15 minute. N-Heptanelayer was decanted and dissolved the solid again in methanol (40 ml).The solvent was distilled out and dried the solid using rotavapor undervacuum. Yield: 7.9 g, HPLC: purity: 98.50%, Chiral purity: 99.50%.

M.P.: 63.6° C.

Example-9 Preparation of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid Potassium salt

In 250 ml round bottom flask, (S,S)(−,−)α-Methyl benzylamine (10.0 g,0.017 mole) salt of2-ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (72 ml) and water (50 ml)was added at RT under N₂ atm., 50% HCl (4.8 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum at50-55° C. Free acid compound was obtained. In a 250 ml three neckedflask, above oily mass (free acid compd.) charged and dissolved inMethanol (39 ml), added potassium tert-butoxide (1.81 g, 0.016 mole)under nitrogen atmosphere and stirred for 30 minutes at RT. The solventwas distilled out using rotavapor. Solid material was obtained.Triturated the solid material with n-Hexane (50 ml) and stirred for 15minute. N-Heptane layer was decanted and dissolved the solid again inmethanol (40 ml). The solvent was distilled out and dried the solidusing rotavapor under vacuum. Yield: 7.6 g (89.2%) HPLC: purity: 98.60%,Chiral purity: 99.56%.

M.P.: 60.4° C.

Example-10 Preparation of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid Zinc salt

In 250 ml round bottom flask, (S,S)-α-Methyl benzylamine (10.0 g, 0.017mole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (72 ml) and water (50 ml)was added at RT under N₂ atm., 50% HCl (4.8 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum 50-55°C. Free acid compound was obtained. In a 250 ml three necked flask,charged above oily mass (free acid compd) and dissolved in Methanol,added a solution of sodium hydroxide (0.929 g in 40 ml water, 0.023mole) and stirred for 15 minute at RT. The reaction mass was cooled at10-15° C. and added a solution of Zinc acetate (2.55 g in 40 ml water,0.011 mole) in 5 minutes. Salt was precipitated out and stirred forfurther 30 minutes. The solid was filtered and washed with water. Driedthe solid under vacuum. Yield: 8.2 g (97.5%), HPLC: purity: 98.97%Chiral purity: 99.55%.

M.P.: 78.5° C.

Example-11 Preparation of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid Lithium salt

In 250 ml round bottom flask, (S,S)-α-Methyl benzylamine (10.0 g, 0.017mole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (72 ml) and water (50 ml)was added at RT under N₂ atm., 50% HCl (4.8 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum at50-55° C. Free acid compound was obtained. In a 250 ml three neckedflask, charged above oily mass (free acid compd.) and dissolved inMethanol (50 ml), added a solution of Lithium hydroxide monohydrate(0.676 g in 4 ml water, 0.016 mole) at RT under nitrogen atmosphere andstirred for 30 minute. The solvent was distilled out using rotavapor.Solid material was obtained. Treated the solid material with n-Hexane(50 ml) and stirred for 15 minute. N-Hexane layer was decanted anddissolved the solid again in methanol (40 ml). The solvent was distilledout and dried the solid using rotavapor under vacuum. Yield: 8.0 g(Quantitative), HPLC: purity: 98.31%, Chiral purity: 99.60%.

M.P.: 85.5° C.

Example-12 Preparation of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid L-arginine salt

In 250 ml round bottom flask, (S,S)-α-methyl benzylamine (10.0 g, 0.017mole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy)}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (72 ml) and water (50 ml)was added at RT under N₂ atm., 50% HCl (4.8 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum at50-55° C. Free acid compound was obtained. In a 250 ml three neckedflask, charged above oily mass (free acid compd) and dissolved inMethanol (50 ml), added a solution of Lithium arginine (2.8 g in 12 mlwater, 0.016 mole) at RT under nitrogen atmosphere and stirred for 30minutes. The solvent was distilled out using rotavapor. Sticky materialwas obtained. Treated the sticky material with n-Hexane (50 ml) andstirred for 15 minutes. N-Hexane layer was decanted and dissolved thesolid again in methanol (40 ml). The solvent was distilled out and driedthe solid using rotavapor under vacuum. Yield: 10.0 g (91.3%), HPLC:purity: 98.67%, Chiral purity: 99.48%.

M.P.: 108.6° C.

Example-13 Preparation of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid Tromethamine salt

In 250 ml round bottom flask, (S,S)-α-Methyl benzylamine (10.0 g, 0.017mole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (85 ml) and water (60 ml)was added at RT under N₂ atm., 50% HCl (5.2 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum. Freeacid compound was obtained. In a 250 ml three necked flask, chargedabove oily mass (free acid compd) and dissolved in Methanol (70 ml),added a solution of Tromethamine (2.34 g in 5 ml water, 0.019 mole) atRT under nitrogen atmosphere and stirred for 30 minutes. The solvent wasdistilled out at rotavapor. Solid material was obtained. Treated thesolid material with n-Hexane (50 ml) and stirred for 15 minute. N-Hexanelayer was decanted and dissolved the solid again in DCM (50 ml). Thesolvent was distilled out and dried the solid using rotavapor undervacuum. Yield: 11.2 g (93.3%), HPLC: purity: 98.40%, Chiral purity:99.43

M.P.: 69.0° C.

Example-14 Preparation of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid L-Lysine salt

In 250 ml round bottom flask, (10.0 g, 0.017 mole) (S,S)-α-Methylbenzylamine salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy)}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (72 ml) and water (50 ml)was added at RT under N₂ atm., 50% HCl (4.8 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum. Freeacid compound was obtained. In a 250 ml three necked flask, chargedabove oily mass (free acid compd.) and dissolved in Methanol (50 ml),added a solution of L-lysine (2.4 g in 8 ml water, 0.016 mole) at RTunder nitrogen atmosphere and stirred for 30 minute. The solvent wasdistilled out at rotavapor. Sticky material was obtained. Treated thesolid-material with n-Hexane (50 ml) and stirred for 15 minutes.N-Hexane layer was decanted and dissolved the solid again in methanol.The solvent was distilled out and dried the solid using rotavapor undervacuum.

Yield: 8.8 g (84.2%), HPLC: purity: 98.91%, Chiral purity: 99.46%.

M.P.: 82.4° C.

Example-15 Preparation of Meglumine salt (i.e. N-Methyl-D-glucamine) of(S) 2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid

In 100 ml round bottom flask, (S,S)-α-Methyl benzylamine (5.0 g, 8.93mmol) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (35 ml) and water (25 ml)was added at RT under N₂ atm. To the suspension 4 ml dilute HCl (2 mlwater+2 ml HCl) solution was added and stirred for 10 minutes. Theorganic layer was separated and washed with water followed by sodiumchloride solution and dried over sodium sulphate. The solvent wasdistilled out using rotavapor under vacuum. Free acid compound wasobtained. To the oily mass (free acid compd.) methanol (25 ml) was addedand stirred to dissolve completely. To the clear reddish solution addeda solution of meglumine (1.6 g dissolved in 10 ml methanol+3 ml water,8.0 mmol) at RT under nitrogen atmosphere. Clear solution was stirredfor 10 minutes under nitrogen atmosphere. The solvent was distilled outusing rotavapor. Sticky material was obtained. Again added 25 mlmethanol to the sticky material. After getting clear solution, solventwas distilled out under vacuum at 45-50° C. Solid was obtained. To thesolid n-Hexane was added and stirred for 15 minutes. N-Hexane wasdecanted. This operation was repeated for another two times to get freeflowing solid. The solid was dried under vacuum at 40° C. for two hours.

Yield: 4.850 g (85.56%), HPLC: purity: 99.15%, Chiral purity: 99.40

M.P.: 72.7° C.

Example-16 Preparation of Benethamine salt (i.e.N-Benzyl-2-phenylethylamine) of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl ethoxyphenyl propionic acid

In 250 ml round bottom flask, (S,S)-α-Methyl benzylamine (0.500 g, 0.892mmole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy)-phenyl)-propionicacid was charged. A solution of Ethyl acetate (3.5 ml) and water (2.5ml) was added at RT under N₂ atm. To the suspension 1 ml dilute HClsolution was added and stirred for 10 minutes to dissolved the solidcompletely. The organic layer was separated and washed with water. Driedthe layer over sodium sulphate and the solvent was distilled out usingrotavapor under vacuum. Free acid compound was obtained. To the oilymass (free acid compound) acetonitrile (2 ml) was added and stirred todissolve completely. To the clear reddish solution added benethamine(0.188 g, 0.982 mmol) at RT under N₂ atmosphere. Hazy solution wasstirred for 3 hours at room temperature under N₂ atmosphere. Kept at0-5° C. overnight. Solvent was distilled out under vacuum. Stickymaterial was obtained. Added n-Hexane (10 ml) and stirred for 15 minute.N-Hexane was decanted. The solid was dried under vacuum at 40° C. for 2hours.

Yield: 0.530 g (91.0%), HPLC: purity: 96.97%, Chiral purity: 100%.

M.P.: 103.1° C.

Example-17 Preparation of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid Piperazine salt

In a 50 ml round bottom flask, (S,S)-α-Methyl benzylamine (10.0 g, 0.017mole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (21 ml) and water (15 ml)was added at RT under N₂ atm., 50% HCl (2.0 ml) solution was added andstirred for 10 minutes. The organic layer was separated and washed withwater. The solvent was distilled out using rotavapor under vacuum at.Free acid compound was obtained. In a 50 ml three necked flask, chargedabove oily mass (free acid compd.) and dissolved in Methanol (25 ml),added a solution of piperazine (0.415 g in 10 ml methanol, 0.004 mole)at RT under nitrogen atmosphere and stirred for 30 minute. The solventwas distilled out under vacuum. Sticky material was obtained. Treatedthe sticky material with n-Hexane (50 ml) and stirred for 15 minute.N-Hexane layer was decanted and dissolved the solid again in acetone (10ml). The solvent was distilled out and dried the solid using rotavaporunder vacuum.

Yield: 2.8 g, HPLC: purity: 97.62%, Chiral purity: 99.62%.

M.P.: 74.5° C.

Example-18 Preparation of Benzylamine salt of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid

In 250 ml round bottom flask, (S,S)(−,−) Methyl benzylamine (10.0 g,17.8 mmole) salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid was charged. A solution of Ethyl acetate (70 ml) and water (50 ml)was added at RT under N₂ atm. To the suspension added 7 ml dilute HClsolution and stirred for 10 minutes. The organic layer was separated andwashed with water. Dried the layer over sodium sulphate. The solvent wasdistilled out using rotavapor under vacuum. Free acid compound wasobtained. To the oily mass (free acid compd.) added acetonitrile (100ml) and stirred to dissolved completely. To the clear reddish solution,added benzylamine (1.9 g, 17.8 mmol) at RT under N₂ atmosphere. Hazysolution was stirred for 1 hour at RT under N₂ atmosphere. Precipitatedsolid was filtered, washed with acetonitrile. Free flowing solid wasdried at 50-55° C.

Yield: 8.5 g (85.56%), HPLC: purity: 97.44%, Chiral purity: ND, M.P.:123° C. ESI-MS: (M+H): 547.4

Example-19 Preparation of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid Dibenzylamine salt

In 250 ml round bottom flask, (S,S)(−,−)α-Methyl benzylamine salt (10.0g, 17.8 mmole) salt of2-ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy)-phenyl)-propionicacid was charged. A solution of Ethyl acetate (70 ml) and water (50 ml)was added at RT under N₂ atm. To the suspension added 7 ml dilute HClsolution and stirred for 10 minutes. The organic layer was separated andwashed with water. Dried the layer over sodium sulphate. The solvent wasdistilled out using rotavapor under vacuum. Free acid compound wasobtained. To the oily mass (free acid compd.) added acetonitrile (35 ml)and stirred to dissolved completely. To the clear reddish solution,added Dibenzylamine (3.5 g, 17.8 mmol) at RT under N₂ atmosphere. Hazysolution was stirred for 1 hour at RT under N₂ atmosphere. Precipitatedsolid was filtered, washed with acetonitrile. Free flowing solid wasdried at 50-55° C.

Yield: 9.2 g (81.0%), HPLC: purity: 97.16%, Chiral purity: 100%

M.P.: 61.5° C.

Example-20 Preparation of Epolamine [i.e2-(pyrrolidine-1-yl)ethanol]salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid

In 50 mL round bottom flask, charged (5.0 g) (8.9 mmole) (S,S) (−,−)α-Methyl benzylamine salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid, 35 mL Ethyl acetate followed by 25 mL water at RT under N₂ atm. Tothe suspension added 3.5 mL dilute HCl solution and stirred for 10mints, to dissolve the solid completely. Layer was separated. Organiclayer was washed with water (20 ml×2) followed by brine wash (20-ml×1),dried over sodium sulphate and distilled out solvent under vacuum. (Freeacid compd obtd as an oil). To the oily mass (free acid compd) addedacetonitrile (30 ml) and stirred to dissolved completely. To the clearreddish solution, added 2-(pyrrolidine-1-yl)ethanol (1.03 g, 8.92 mmol)at RT under N₂ atm., hazy solution was stirred for 3 hours at roomtemperature under N₂ atmosphere. Kept at 0-5° C. for overnight. Solid donot precipitate, distilled out solvent under vacuum. Sticky materialobtained. Triturated with n-Hexane (2×50 ml), decanted n-Hexane layer,then again dissolved in methylene chloride (10 ml), added into n-heptaneunder stirring, solid do not precipitate, distilled out solvent undervacuum at 50-55° C.

Yield: 5.1 g (quantitative), HPLC: purity: 96.76%, Chiral purity: 99.82%

Example-21 Preparation of Imidazole [i.e 1H-Imidazole]salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid

In 100 ml round bottom flask, charged (5.0 g) (8.92 mmole) (S,S) (−,−)α-Methyl benzylamine salt of2-ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy)-phenyl)-propionicacid, 35 ml Ethyl acetate followed by 25 ml water at RT under N₂ atm. Tothe suspension added 3.4 ml dilute HCl solution and stirred for 10mints, to dissolve the solid completely layer was separated. Organiclayer was washed with water (20 ml×2), dried over sodium sulphate anddistilled out solvent under vacuum. (Free acid compd. obtd. as anoil-4.2 g). To the oily mass (free acid compd) added acetonitrile (30ml) and stirred to dissolved completely. To the clear reddish solution,added 1H imidazole (0.608 g, 8.92 mmol) dissolved in 10 ml acetonitrileat RT under N₂ atm., hazy solution was stirred for 2 hours at roomtemperature under N₂ atmosphere. Kept at 0-5° C. for overnight. Solventdistilled off completely under vacuum, added n-hexane (2×50 ml) andstirred for 10 minutes, decanted n-hexane, oily material dissolved inmethylene chloride (10 ml), dumped into 100 ml n-heptane under stirring,sticky semi solid was scratched and stirred for 1 hour at roomtemperature. Free flowing solid was filtered, washed with n-heptane(2×20 ml), suck dried. Solid was dried under vacuum at 50-55° C.

Yield: 4.2 g (93%), HPLC: purity: 97.00%, Chiral purity: 100%, M.P.:106.6° C.

Example-22 Preparation of Triethanolamine salt of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid

In 250 ml round bottom flask, charged (5.0 g) (8.92 mmole) (S,S) (−,−)α-Methyl benzylamine of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid, 35 ml Ethyl acetate followed by 25 ml water at RT under N₂ atm. Tothe suspension added 3.5 ml dilute HCl solution and stirred for 10 mintsto dissolve the solid completely. Layer was separated. Organic layer waswashed with water (30 ml×2) followed by brine wash (30 ml×1). Dried oversodium sulphate and distilled out solvent under vacuum. (Free acidcompound obtained as oil-4.3 g). To the oily mass (free acid compd)added acetonitrile (30 ml) and stirred to dissolved completely. To theclear reddish solution, added triethanol amine (1.34 g, 8.92 mmol)diluted with 10 ml acetonitrile at RT under N₂ atm., clear solution wasstirred for 2-3 hours at room temperature under N₂ atmosphere. Kept at0-5° C. for overnight Solvent distilled off completely under vacuum,added n-hexane (10 ml) and stirred for 10 minutes, distilled offn-hexane, oily material dissolved in methylene chloride (10 ml), dumpedinto 100 ml n-heptane under stirring, sticky oily material was scratchedand stirred for 2 hours at room temperature. Solid was not precipitated,distilled out solvent under vacuum at 50-55° C. Oily/pasty material wasobtained.

Yield: 5.8 g (Quantitative yield), HPLC: purity: 96.80%, Chiral purity:100%

Example-23 Preparation of Ethanolamine salt of (S)2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid

In 50 ml round bottom flask, charged (5.0 g) (8.92 mmole) (S,S) (−,−)α-Methyl benzylamine salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid, 35 ml Ethyl acetate followed by 25 ml water at RT under N₂ atm. Tothe suspension added 3.5 ml dilute HCl solution (1.7 ml HCl+1.7 mlwater) and stirred for 10 mints, to dissolve the solid completely.

Layer was separated. Organic layer was washed with water (20 ml×2)followed by brine wash (20 ml×1), dried over sodium sulphate anddistilled out solvent under vacuum. (Free acid compd obtd as an oil). Tothe oily mass (free acid compd) added acetonitrile (30 ml) and stirredto dissolved completely. To the clear reddish solution, addedmonoethanol amine (0.546 g, 8.92 mmol) diluted with 10 ml acetonitrileat RT under N₂ atm., hazy solution was stirred for 2 hours at roomtemperature under N₂ atmosphere. Solvent distilled off completely undervacuum, oily material dissolved in methylene chloride (10 ml), dumpedinto 100 ml n-heptane under stirring, sticky oily material was scratchedand stirred for 1 hour at room temperature. Solid was not precipitated,distilled out solvent under vacuum at 50-60° C. Oily/pasty materialobtained.

Yield: 4.7 g (Quantitative yield), HPLC purity: 97.06%, Chiral purity:100%

Example-24 Preparation of Choline salt (i.e2-Hydroxy-N,N,N-trimethylethanamiaiam hydroxide or choline hydroxide) of(S) 2-Ethoxy-3-(4-(2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl ethoxyphenyl propionic acid

In 50 ml round bottom flask, charged (3.0 g) (5.35 mmole) (S,S) (−,−)α-Methyl benzylamine salt of2-ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy)-phenyl)-propionicacid, 21 ml Ethyl acetate followed by 15 ml water at RT under N₂ atm. Tothe suspension added 2 ml dilute HCl solution (1 ml HCl+1 ml water) andstirred for 10 mints to dissolve the solid completely. The layer wasseparated. Organic layer was washed with water (15 ml×2), dried oversodium sulphate and distilled out solvent under vacuum. (Free acidcompd. obtd as an oil-2.4 g). To the oily mass (free acid compd-2.4 g)added acetonitrile (15 ml) and stirred to dissolved completely. To theclear reddish solution, added 3.24 ml of 20% choline hydroxide solution(0.649 g, 5.35 mmol) at RT under N₂ atm., hazy solution was stirred for2 hours at room temperature under N₂ atmosphere. Kept in freeze forovernight. Solvent distilled off completely under vacuum, gummy materialobtained, added n-hexane (30 ml), stirred & decanted, againn addedn-hexane (30 ml), stirred & decanted, dissolved gummy material inmethylene chloride (10 ml), dumped into 100 ml n-heptane under stirring,sticy oily material was scratched and stirred for 2 hour at roomtemperature. Solid was not precipitated, distilled out solvent undervacuum at 50-60° C.

Yield: 2.9 g (100%), HPLC: purity: 94.01%, Chiral purity: 100%

Example-25 Preparation of (s)-□-phenyl ethylamine salt(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

In a 50 mL flask (S)-□-phenyl ethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (5 g), water (5 mL) and isopropyl acetate (25 mL) were mixed underN₂ atmosphere at 25-30° C. To the mixture 35% Conc. HCl (1.11 mL) wasadded slowly with stirring. It was stirred vigorously upto soliddissolved completely at 25-30° C. It was transferred into the separatingfunnel. The organic layer was collected and washed with water (2.5 mL).It was dried over anhydrous sodium sulfate. The solvent was distilledout under reduced pressure on Buchi Rota vapour to obtain thick liquid.(Wt.: 3.9 g)

Example-26 Preparation of free acid of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

In a 25 mL flask Magnesium salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (2 g), water (2 mL) and ethyl acetate (10 mL) were mixed under N₂atmosphere at 25-30° C. To the mixture 35% Cone. HCl (0.55 mL) was addedslowly with stirring. It was stirred vigorously till solid dissolvedcompletely at 25-30° C. It was transferred into the separating funnel.The organic layer was collected and washed with water (2×2 mL). It wasdried over anhydrous sodium sulfate. The solvent was distilled out underreduced pressure on Buchi Rota vapour to obtain thick liquid free acid.(Wt.: 1.9 g)

Example-27 Preparation of Dicyclohexylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (3.9 g) was freshly prepared and dissolved in acetonitrile (25 mL)at 25-30° C. To the clear solution dicyclohexylamine (1.61 g) was addedwith stirring and stirred for 24 h at 25-30° C., solid salt wasprecipitated. It was diluted with acetonitrile (15 mL). Thedicyclohexylamine salt was filtered and washed with acetonitrile (3×5mL). It was dried under vacuum in the flask. Wt.: 2.3 g, % Y-41.6%, %Purity by HPLC-98.9%, m.p. 98-100° C.

¹H NMR (CDCl₃, 300 MHz): δ 1.08 (t, 3H), 1.11-1.1.24 (m, 5H), 1.40-1.44(m, 5H), 1.60 (m, 2H), 1.75-1.79 (m, 4H), 1.97-2.00 (m, 4H), 2.36 (s,3H), 2.50 (s, 3H), 2.83-2.95 (m, 4H), 3.27 (m, 1H), 3.62 (m, 1H), 3.76(m, 1H), 3.89 (t, 2H), 4.24 (t, 2H), 5.94 (d, 1H, J=3.3 Hz), 6.08 (d,1H, J=3.3 Hz), 6.59 (d, 2H, J=8.7 Hz), 7.15 (d, 2H, J=8.7 Hz), 7.26-7.34(m, 4H).

Example-28 Preparation of (S)-α-naphthylethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (1.95 g) was freshly prepared and dissolved in acetonitrile (20 mL)at 25-30° C. To the clear solution (S)-α-naphthylethylamine (0.763 g)was added with stirring, solid salt was precipitated. It was heated toreflux temperature and stirred for 10 min. at 80-82° C. It was cooled to25-30° C. and stirred for 15 min. at 25-30° C. The(S)-α-naphthylethylamine salt was filtered and washed with acetonitrile(3×10 mL). It was dried under vacuum in the flask. Wt.: 2.2 g, %Y-80.9%, % Purity by HPLC-98.9%, m.p. 153-154° C.

¹H NMR (CDCl₃, 300 MHz): δ 0.95 (t, 3H), 1.65 (d, 3H), 2.33 (s, 3H),2.48 (s, 3H), 2.56 (m, 1H), 2.71 (m, 1H), 3.03 (m, 1H), 3.37 (m, 1H),3.62 (m, 1H), 3.83 (t, 2H), 4.22 (t, 2H), 5.14 (q, 1H), 5.94 (d, 1H,J=3.3 Hz), 6.08 (d, 1H, J=3.3 Hz), 6.48 (d, 2H, J=8.7 Hz), 6.86 (d, 2H,J=8.7 Hz), 7.23-7.31 (m, 4H), 7.44-7.53 (m, 3H), 7.74 (m, 2H), 7.82 (m,1H), 7.98 (m, 1H).

m/z: 440 (M+H)+, 100%

Example-29 Preparation of (S)-α-3-methoxyphenylethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (1.56 g) was freshly prepared and dissolved in acetonitrile (15 mL)at 25-30° C. To the clear solution (S)-α-3-methoxyphenylethylamine(0.540 g) was added with stirring and stirred for 30 min. at 25-30° C.,solid salt was precipitated. The (S)-α-3-methoxyphenylethylamine saltwas filtered and washed with acetonitrile (3×3 mL). It was dried undervacuum in the flask.

Wt.: 1.7 g, % Y-80.9%, % Purity by HPLC-98.5%, m.p. 139-141° C.

¹H NMR (CDCl₃, 300 MHz): δ 0.98 (t, 3H), 1.51 (d, 3H), 2.35 (s, 3H),2.49 (s, 3H), 2.61 (m, 1H), 2.76 (m, 1H), 3.06 (m, 1H), 3.40 (m, 1H),3.63 (m, 1H), 3.71 (s, 3H), 3.89 (t, 2H), 4.15 (q, 1H), 4.25 (t, 2H),5.37 (s, br, —NH₂, 2H), 5.94 (d, 1H, J=3.3 Hz), 6.07 (d, 1H, J=3.3 Hz),6.55 (d, 2H, J=8.7 Hz), 6.77 (m, 1H), 6.99 (m, 4H), 7.18-7.33 (m, 5H).

m/z: 440 (M+H)⁺, 100%

The following salts were prepared following processes similar to thoseabove along with suitable modifications which are within the scope of askilled person.

Example-30 Preparation of (S)-α-4-mehoxyphenylethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (1.56 g) was reacted with (S)-α-4-methoxyphenylethylamine (0.555 g)in acetonitrile.

Wt.: 1.9 g, % Y-90.4%, % Purity by HPLC-98.6%, m.p. 145-147° C.

¹H NMR (CDCl₃, 300 MHz): δ 0.96 (t, 3H), 1.51 (d, 3H), 2.35 (s, 3H),2.49 (s, 3H), 2.58 (m, 1H), 2.71 (m, 1H), 2.98 (m, 1H), 3.36 (m, 1H),3.55 (m, 1H), 3.66 (t, 3H), 3.89 (t, 2H), 4.14 (q, 1H), 4.25 (m, 2H),5.80 (s, br, 2H, —NH₂), 5.94 (d, 1H, J=3.3 Hz), 6.08 (d, 1H, J=3.3 Hz),6.56 (d, 2H, J=8.7 Hz), 6.76 (d, 2H, J=8.7 Hz), 6.98 (d, 2H), 7.24-7.36(m, 6H) m/z: 440 (M+H)⁺, 100%

Example-31 Preparation of (S)-α-4-chlorophenylethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (1.56 g) was reacted with (S)-α-4-chlorophenylethylamine (0.555 g)in acetonitrile.

Wt: 1.9 g, % Y-89.6%, % Purity by HPLC-98.5%, m.p. 161-163° C.

¹H NMR (CDCl₃, 300 MHz): δ 0.99 (t, 3H), 1.47 (d, 3H), 2.35 (s, 3H),2.49 (s, 3H), 2.68 (m, 1H), 2.78 (m, 1H), 3.10 (m, 1H), 3.35 (m, 1H),3.66 (dd, 1H), 3.90 (t, 2H), 4.10 (q, 1H), 4.26 (t, 2H), 4.74 (s,br, 2H,—NH₂), 5.94 (d, 1H, J=3.3 Hz), 6.08 (d, 1H, J=3.3 Hz), 6.57 (d, 2H,J=8.7 Hz), 6.99 (d, 2H), 7.24-7.35 (m, 8H).

m/z: 440 (M+H)⁺, 100%

Example-32 Preparation of (S)-α-4-methylphenylethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (1.56 g) was reacted with (S)-α-)-α-4-methylphenylethylamine (0.482g) in acetonitrile.

Wt.: 1.85 g, % Y-90.2%, % Purity by HPLC-98.6%, m.p. 152-154° C.

¹H NMR (CDCl₃, 300 MHz): δ 0.97 (t, 3H), 1.51 (d, 3H), 2.24 (s, 3H),2.35 (s, 3H), 2.49 (s, 3H), 2.58 (m, 1H), 2.71 (m, 1H), 2.99 (m, 1H),3.36 (m, 1H), 3.55 (dd, 1H), 3.90 (t, 2H), 4.17 (q, 1H), 4.25 (m, 2H),5.60 (s, br, —NH₂), 5.94 (d, 1H, J=3.3 Hz), 6.08 (d, 1H, J=3.3 Hz), 6.56(d, 2H, J=8.7 Hz), 6.97 (d, 2H, J=8.7 Hz), 7.06 (d, 2H), 7.24-7.33 (m,6H).

m/z: 440 (M+H)⁺, 100%

Example-33 Preparation of 1-Adamantanamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (1.95 g) was freshly prepared from Mg salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (2 g) and dissolved in ethylacetate (10 mL) at 25-30° C. To theclear solution I-adamantanamine (0.670 g) was added with stirring andstirred for 1.5 h at 25-30° C., solid salt was precipitated. It wasdiluted with 40 mL ethyl acetate. The 1-adamantanamine salt was filteredand washed with ethylacetate (2×10 mL). It was dried under vacuum in theflask.

Wt.: 1.9 g, % Y-72.0%, % Purity by HPLC-98.7%

Example-34 Preparation of 1-adamantanemethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

1-Adamantanemethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid was prepared following above procedure from(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid (0.975 g)

Wt.: 0.7 g, % Y-52.0%, % Purity by HPLC-99.1%

Example-35 Preparation of Diphenylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid (3.9 g) was freshly prepared and dissolved in acetonitrile (40 mL)at 25-30° C. To the clear solution diphenylamine (1.5 g) was added withstirring and stirred for 2 h at 25-30° C. It was cooled to 0-5° C. andstirred for 20 h. Solid was not precipitated. The solvent was distilledout under reduced pressure on Buchi Rota vapour to obtain thick liquid.

Wt.: 5.1 g, % Y-94.0%.

Example-36 Preparation of Diethylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid (1.95 g) was freshly prepared and dissolved in isopropylacetate (20mL) at 25-30° C. To the clear solution diethylamine (0.46 g) was addedwith stirring and stirred for 18 h at 25-30° C., solid did notprecipitated. The solvent was distilled out under reduced pressure onBuchi Rota vapour to obtain thick liquid.

Wt.: 2.2 g, % Y-96.9%.

Example-37 Preparation of α-naphthylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid (3.9 g) was freshly prepared and dissolved in acetonitrile (40 mL)at 25-30° C. To the clear solution α-naphthylamine (1.27 g) was addedwith stirring and stirred for 2 h at 25-30° C. It was cooled to 0-5° C.and stirred for 20 h, solid was not precipitated. The solvent wasdistilled out under reduced pressure on Buchi Rota vapour to obtainthick liquid.

Wt.: 4.8 g, % Y-92.8%.

Example-38 Preparation of O-phenylenediamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid (3.9 g) was freshly prepared and dissolved in acetonitrile (20 mL)at 25-30° C. To the clear solution O-phenylenediamine (0.970 g) wasadded with stirring and stirred for 3 h at 25-30° C. It was cooled to0-5° C. and stirred for 20 h, solid was not precipitated. The solventwas distilled out under reduced pressure on Buchi Rota vapour to obtainthick liquid.

Wt.: 4.6 g, % Y-94.0%.

Example-39 Preparation of 1,3-diaminopropane salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid (1.95 g) was freshly prepared and dissolved in acetonitrile (10 mL)at 25-30° C. To the clear solution 1,3-diaminopropane (0.330 g) wasadded with stirring and stirred for 1 h at 25-30° C. It was cooled to0-5° C. and stirred for 1 h, liquid separated. The solvent was distilledout under reduced pressure on Buchi Rota vapour to obtain thick liquid.

Wt.: 1.95 g, % Y-85.1%.

Example-40 Preparation of tritylamine salt of(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]-ethoxy}-phenyl)propanoicacid

(S)-2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)pyrrol-1-yl]ethoxy}phenyl)propanoicacid (1.95 g) was freshly prepared and dissolved in ethylacetate (10 mL)at 25-30° C. To the clear solution tritylamine (1.15 g) was added withstirring and stirred for 24 h at 25-30° C., solid did not precipitate.The solvent was distilled out under reduced pressure on Buchi Rotavapour to obtain thick liquid.

Wt.: 3.0 g, % Y-96.7%.

Similarly the following salts of compound of formula (I) with followingbase were also prepared. Cinchonine, Cinchonidine, (−)-Quinine,Ethylenediamine, Morpholine-4-(2-hydroxyethyl), N,N-Diethylethanolamine, Deanol (i.e. N,N-Dimethyl ethanolamine), Hydrabmine,Betaine, Ammonia.

The salts of the invention can be used either for the chemical & chiralpurification of the compounds of formula (I) or whenever possible, aspharmaceutically acceptable compounds. When used as pharmaceuticallyacceptable compounds, the compound of formula (I) is formulated intosuitable pharmaceutical formulations using suitable binders and otherexcipients. Such pharmaceutical compositions may be suitable for thetreatment of suitable mammals when such mammals are in need of suchtreatment. The dosing regimen will be decided based on the therapeuticintervention, the species being treated and the severity of the disease.A skilled person can decide on these based on his knowledge andexpertise.

Biological studies:

Comparative Efficacy Study of (Triglyceride Lowering Effects) of VariousSalts of Compound (Ia) in Swiss Albino Mice.

The in-vivo efficacy of compound (I) was evaluated in Swiss albino mice.Anti-dyslipidemic drugs have been reported to lower circulating levelsof triglyceride in Swiss albino mice through their effect on genesinvolved in the peroxisomal fatty acid beta oxidation via PPAR alphaagonism. Therefore, this species is preferred for evaluation of theirefficacy in lowering circulating triglyceride (TG) levels.

In this experiment, six to eight week old male Swiss albino mice wereissued and kept for acclimatization. Near the end of the acclimatizationperiod, animals judged to be suitable for testing were bled under lightether anesthesia and serum samples were analyzed for Serum triglyceridelevels. Animals were selected according to triglyceride levels in therange of 70-180 mg/dl and divided into groups of 6 animals each suchthat the average TG levels of animals in each group were notsignificantly different from the others.

Test compound was formulated at specified doses in vehicle (10%Polyethylene glycol (PEG) 400+90% of 0.5% Sodium Carboxy methylcellulose). The animals were dosed orally, once daily in the morningduring six days, starting from next day of grouping with vehicle or testcompound. The animals were weighed prior to dosing, and based on theseweights; the volume of administration was calculated. The volume offormulation administered to each mouse was 10 ml/kg body weight.

On day 6, one hr after the dose administration, blood (0.25 ml) wascollected from retro-orbital sinus of the anaesthetized animals. Serumwas separated by centrifugation. Serum was analyzed for triglyceridelevels. Analysis for serum triglyceride levels was performed usingSpectrophotometer and commercially available kit. Calculations fordetermination of % change and % reduction in serum TG levels wereperformed using MS Excels sheet.

TABLE NO. 2 Treatment groups and dose levels Dose Number of GroupTreatment (mg/kg) Animals 1 Vehicle Control [10% PEG400 + 90% 0 mg/kg 6Na-CMC(0.5%)] 2 Magnesium salt of formula (I) at 1.0252 mg/kg 1 mg/kg 6equivalent to 1 mg/kg of corresponding free acid 3 Calcium Salt ofFormula (I) at 1.0437 mg/kg 1 mg/kg 6 equivalent to 1 mg/kg ofcorresponding free acid 4 Sodium Salt of Formula (I) at 1.0498 mg/kg 1mg/kg 6 equivalent to 1 mg/kg of corresponding free acid 5 PotassiumSalt of Formula (I) at 1.0818 mg/kg 1 mg/kg 6 equivalent to 1 mg/kg ofcorresponding free acid 6 Zinc Salt of Formula (I) at 1.0664 mg/kg 1mg/kg 6 equivalent to 1 mg/kg of corresponding free acid

TABLE NO. 3 Effect on triglyceride levels after 6 days of treatment.Serum triglycerides (mg/dl) % Change % Change Compound Day 0 Day 6 Day 6vs Day 0 Vs control Vehicle Control 119.7 ± 10.2 118.2 ± 7.8   1.4 ± 9.6[10% PEG400 + 90% Na-CMC(0.5%)] Magnesium salt of 117.7 ± 13.8 38.2 ±4.1 −66.5 ± 4.1 −66.1 ± 4.2 formula (I) at 1.0252 mg/kg equivalent to 1mg/kg of corresponding free acid Calcium Salt of 119.2 ± 11.1 38.8 ± 3.2−66.8 ± 2.5 −66.4 ± 2.5 Formula (I) at 1.0437 mg/kg equivalent to 1mg/kg of corresponding free acid Sodium Salt of 119.4 ± 11.9 41.6 ± 1.8−63.5 ± 3.6 −63.1 ± 3.6 Formula (I) at 1.0498 mg/kg equivalent to 1mg/kg of corresponding free acid Potassium Salt of 119.0 ± 9.2  44.8 ±4.6 −60.7 ± 6.0 −60.2 ± 6.0 formula (I) at 1.0818 mg/kg equivalent to 1mg/kg of corresponding free acid Zinc Salt of Formula 119.1 ± 7.6  68.6± 7.5 −42.7 ± 4.2 −42.0 ± 4.2 (I) at 1.0664 mg/kg equivalent to 1 mg/kgof corresponding free acid

TABLE NO. 4 Treatment groups and dose levels Number Dose of GroupTreatment (mg/kg) Animals 1 Vehicle Control [10% PEG400 + 90% 0 mg/kg 6Na-CMC(0.5%)] 2 Mg salt of formula (I) at 1.0252 mg/kg 1 mg/kg 6equivalent to 1 mg/kg of corresponding free acid 3 L-arginine salt ofFormula (I)) at 1.284 1 mg/kg 6 mg/kg equivalent to 1 mg/kg ofcorresponding free acid 4 Tromethamine salt of Formula (I) at 1.2162 1mg/kg 6 mg/kg equivalent to 1 mg/kg of corresponding free acid 5L-lysine salt of Formula (I) at 1.2498 mg/kg 1 mg/kg 6 equivalent to 1mg/kg of corresponding free acid 6 Piperazine salt of Formula (I) at1.1638 1 mg/kg 6 mg/kg equivalent to 1 mg/kg of corresponding free acid7 Dibenzylamine salt of Formula (I) at 1.3097 1 mg/kg 6 mg/kg equivalentto 1 mg/kg of corresponding free acid 8 Benethamine salt of Formula (I)at 1.3246 1 mg/kg 6 mg/kg equivalent to 1 mg/kg of corresponding freeacid 9 Benzylamine salt of Formula (I) at 1.196 1 mg/kg 6 mg/kgequivalent to 1 mg/kg of corresponding free acid

TABLE NO. 5 Effect on triglyceride levels after 6 days of treatment.Serum triglycerides (mg/dl) % Change % Change Compound Day 0 Day 6 Day 6vs Day 0 Vs control Vehicle Control 91.6 ± 5.0 96.8 ± 8.3  6.7 ± 9.6[10% PEG400 + 90% Na- CMC(0.5%)], po Mg salt of formula (I) at 91.4 ±8.8 37.4 ± 3.2 −58.3 ± 3.7 −60.5 ± 3.5 1.0252 mg/kg equivalent to 1mg/kg of corresponding free acid L-arginine salt of Formula 91.6 ± 7.937.2 ± 2.1 −58.0 ± 4.2 −60.2 ± 3.9 (I) at 1.284 mg/kg equivalent to 1mg/kg of corresponding free acid Tromethamine salt of 91.2 ± 7.8 47.9 ±4.9 −43.9 ± 9.0 −46.9 ± 8.5 Formula (I) at 1.2162 mg/kg equivalent to 1mg/kg of corresponding free acid L-lysine salt of Formula (I) 91.2 ± 6.848.7 ± 3.3 −45.8 ± 3.8 −48.7± at 1.2498 mg/kg equivalent to 1 mg/kg ofcorresponding free acid Piperazine salt of Formula 91.0 ± 6.5 44.1 ± 1.9−50.2 ± 4.0 −52.8± (I) at 1.1638 mg/kg equivalent to 1 mg/kg ofcorresponding free acid Dibenzylamine salt of 91.0 ± 6.7 42.1 ± 2.9−51.8 ± 6.1 −54.4± Formula (I) at 1.3097 mg/kg equivalent to 1 mg/kg ofcorresponding free acid Benethamine salt of Formula 91.6 ± 6.4 56.0 ±4.9 −38.3 ± 5.7 −41.6± (I) at 1.3246 mg/kg equivalent to 1 mg/kg ofcorresponding free acid Benzylamine salt of Formula  91.1 ± 10.8 36.5 ±4.0 −57.9 ± 6.2 −60.2± (I) at 1.196 mg/kg equivalent to 1 mg/kg ofcorresponding free acid

TABLE NO. 6 Treatment groups and dose levels Dose Number of GroupTreatment (mg/kg) Animals 1 Vehicle Control [10% PEG400 + 90% 0 mg/kg 6Na-CMC(0.5%)] 2 Mg salt of compound of formula (I) at 1 mg/kg 6 1.0252mg/kg, po equivalent to 1 mg/kg of corresponding free acid 3 Compound laat 1.3075 mg/kg, po 1 mg/kg 6 equivalent to 1 mg/kg of correspondingfree acid 4 Triethanolamine salt of Formula (I) at 1.25 1 mg/kg 6 mg/kg,po equivalent to 1 mg/kg of corresponding free acid 5 Ethanolamine saltof Formula (I) at 1.12 1 mg/kg 6 mg/kg, po equivalent to 1 mg/kg ofcorresponding free acid 6 Epolamine salt of Formula (I) at 1.2 1 mg/kg 6mg/kg, po equivalent to 1 mg/kg of corresponding free acid 7 Imidazolesalt of Formula (I) at 1.13 1 mg/kg 6 mg/kg, po equivalent to 1 mg/kg ofcorresponding free acid 8 Choline salt of Formula (I) at 1.22 1 mg/kg 6mg/kg, po equivalent to 1 mg/kg of corresponding free acid

TABLE NO. 7 Effect on triglyceride levels after 6 days of treatmentSerum triglycerides (mg/dl) % Change % Change Compound Day 0 Day 6 Day 6vs Day 0 Vs control Vehicle Control 111.0 ± 8.0 99.6 ± 8.8  −9.8 ± 5.8[10% PEG400 + 90% Na- CMC(0.5%)] Mg salt of formula (I) at 111.1 ± 8.436.9 ± 2.9 −65.5 ± 4.2 −61.5 ± 4.6 1.0252 mg/kg, po equivalent to 1mg/kg of corresponding free acid Maglumine salt of Formula 110.6 ± 9.132.4 ± 1.5 −70.0 ± 2.2 −66.5 ± 2.5 (I) at 1.3075 mg/kg, po equivalent to1 mg/kg of corresponding free acid Triethanolamine salt of 110.3 ± 7.533.0 ± 2.6 −68.9 ± 3.9 −65.3 ± 4.4 Formula (I) at 1.25 mg/kg, poequivalent to 1 mg/kg of corresponding free acid Ethanolamine salt ofFormula 110.5 ± 7.5 42.5 ± 3.6 −61.3 ± 3.2 −56.9 ± 3.6 (I) at 1.12mg/kg, po equivalent to 1 mg/kg of corresponding free acid Epolaminesalt of Formula 110.5 ± 7.5 39.6 ± 3.8 −64.2 ± 2.3 −60.1 ± 2.6 (I) at1.2 mg/kg, po equivalent to 1 mg/kg of corresponding free acid Imidazolesalt of Formula (I) 110.5 ± 7.2 43.5 ± 2.5 −59.8 ± 3.2 −55.3 ± 3.6 at1.13 mg/kg, po equivalent to 1 mg/kg of corresponding free acid Cholinesalt of Formula (I) at 110.9 ± 7.7 47.6 ± 3.4 −56.0 ± 4.3 −51.0 ± 4.81.22 mg/kg, po equivalent to 1 mg/kg of corresponding free acid

Efficacy Studies of Synergistic Combination Containing Compound ofFormula (I) where M⁺ is Mg⁺²:

I) Antidiabetic Activity of Compound (Ia), Sitagliptin (a DipeptidylPeptidase IV Inhibitor) and Co-Administration of Compound(Ia)+Sitagliptin in db/db Mice Model

II) Antidiabetic Activity of Compound (Ia), Metformin andCo-Administration of Compound (Ia)+Metformin in db/db Mice Model

The in-vivo efficacy for antidiabetic activity was done for (I) and (II)individually with db/db mice of 13-14 weeks of age. Generally obese anddiabetic animals are insulin resistant and have abnormalities in glucoseand lipid metabolism. The development of diabetes and beta celldysfunction in these animal models closely parallels the pathophysiologyof the disease condition in humans. The db/db mouse exhibits an initialphase of hyperinsulinemia, hyperphagia and obesity. They progressivelydevelop into insulinopenia with age, a feature commonly observed in laststage of type-2 diabetes, when blood sugar levels are insufficientlycontrolled. Therefore, this genetic model proves to be suitable forpredicting the likely therapeutic benefit of novel euglycemic agents inhuman type-2 diabetes.

In this experiment, 48 female db/db mice of 13-14 week were divided into6 groups and as per their day 0 serum glucose levels per Table no. 8.Vehicle Control group was treated with vehicle (5% Polyethylene glycol(PEG) 400+5% Tween 80+90% of 0.5% Sodium Carboxy methyl cellulose) andother five groups with treatment mentioned in Table no. 8. The treatmentwas given oral gavages once daily for 14 days.

TABLE NO. 8 Dosage Number of Sr. no Treatment Group Level Sex Animals 1Vehicle Control 0 mg/kg Female 8 2 Compound (Ia) 0.1 mg/kg   Female 8 3Sitagliptin 3 mg/kg Female 8 4 Compound (Ia) 0.1 mg/kg + Female 8 (0.1mg/kg) + 3 mg/kg Sitagliptin (3 mg/kg) 5 Metformin 150 mg/kg  Female 8 6Compound (Ia) 0.1 mg/kg + Female 8 (0.1 mg/kg) + 150 mg/kg  Metformin(150 mg/kg) 

The blood was collected by retro-orbital sinus puncture method underlight ether anesthesia on day 0 (pretreatment) and on day 12 fornon-fasting glucose analysis and oral glucose tolerance test (OGTT) wasperformed on day 14 of the treatment in fasting condition. After bloodcollection serum was separated and analyzed glucose analysis usingSpectra max and commercial diagnostic kits. The OGTT procedure followedis as given below—

Oral Glucose Tolerance Test (OGTT):

One day before OGTT all animals were kept on overnight fasting and onthe day of OGTT, day 14 of the treatment each animal was dosed with asingle dose of vehicle/test compounds administered orally and 60-minpost dosing blood was collected (0 min) and glucose load (2 gm/kg/10 ml)administered per orally. Blood was then collected at time pointscorresponding to 30, 60 and 120 min after glucose load administration.Serum was separated and analyzed for glucose using Spectra max andcommercial diagnostic kits.

The calculation for antidiabetic activity i.e. effect on serum glucoseand improvement in glucose Area Under Curve (AUC) was calculated usingthe excel sheet and Graph Pad Prism software. The day 0 and day 12glucose values of vehicle control and treatment were considered forcalculating the effect on serum glucose. The effect on glucose AUC wascalculated using the OGTT glucose levels at different time points andthe improvement in glucose AUC were the reduction in glucose AUC afterthe treatment during OGTT.

The effect on serum glucose levels in different treatment groups and %change Vs. Vehicle Control is depicted in Table no. 9.

TABLE NO. 9 % Change Serum Glucose (mg/dl) Day 12 vs % Change CompoundDay 0 Day 12 Day 0 Vs control Vehicle control 406.7 ± 27.1 405.7 ± 39.1 1.3 ± 9.4 Compound 407.8 ± 28.9 276.9 ± 39.4 −33.0 ± 5.9 −32.8 ± 6.0(Ia)- 0.1 mg/kg, p.o. Sitagliptin- 407.9 ± 34.5 485.3 ± 50.5  18.3 ± 5.0 18.5 ± 5.0 3 mg/kg, p.o. Compound 393.2 ± 27.9 161.5 ± 20.1 −59.1 ± 3.1−59.0 ± 3.1 (Ia)- 0.1 mg/kg, p.o. + Sitagliptin- 3 mg/kg, p.o.

The treatment with Compound (Ia) at 0.1 mg/kg showed 32.8% reduction inserum glucose and Sitagliptin (dipeptidyl peptidase IV inhibitor)treatment does not show any reduction rather it has shown slightincrease in serum glucose whereas co-administration of Compound (Ia) at0.1 mg/kg+Sitagliptin (dipeptidyl peptidase IV inhibitor) at 3 mg/kg hasshown synergistically 59% reduction in serum glucose.

The effect on glucose area under curve in OGTT in different treatmentgroups and % change Vs Vehicle Control is depicted in Table no. 10

TABLE NO. 10 % improvement AUC Glucose in AUC Compounds (mg/dl · min)glucose vs control Vehicle control 56845.5 ± 4313.6 Compound (Ia)- 0.1mg/kg, p.o. 51757.3 ± 3366.3  9.0 ± 5.9 Sitagliptin- 3 mg/kg, p.o.64655.9 ± 6300.6 −13.7 ± 11.1 Compound (Ia)- 0.1 mg/kg, p.o. + 38516.4 ±4515.3 32.2 ± 7.9 Sitagliptin- 3 mg/kg, p.o.

In oral glucose tolerance test, Compound (Ia) at 0.1 mg/kg showed only9% improvement in glucose area under curve and Sitagliptin treatmentdoes not show any improvement rather it has shown slight increase inglucose intolerance whereas co-administration of Compound (Ia) at 0.1mg/kg+Sitagliptin at 3 mg/kg has shown synergistically 32.2% improvementin glucose area under curve during the OGTT done after 14 days oftreatment, so it indicates that co-administration of Compound (Ia) andSitagliptin has shown synergistic activity in both the antidiabeticparameters evaluated i.e. serum glucose and oral glucose tolerance test.

The effect on serum glucose levels in different treatment groups and %change Vs Vehicle Control is depicted in Table no. 11.

TABLE NO. 11 % Change Serum Glucose (mg/dl) Day 12 vs % Change CompoundDay 0 Day 12 Day 0 Vs control Vehicle control 406.7 ± 27.1 405.7 ± 39.1 1.3 ± 9.4 Compound 407.8 ± 28.9 276.9 ± 39.4 −33.0 ± 5.9 −32.8 ± 6.0(Ia)- 0.1 mg/kg p.o. Compound 405.8 ± 26.9 217.5 ± 26.4 −45.5 ± 4.1−45.4 ± 4.4 (Ia)- 0.1 mg/kg, p.o. + Metfomin- 150 mg/kg, p.o.

The treatment with Compound (Ia) at 0.1 mg/kg showed 32.8% reduction in,serum glucose whereas co-administration of Compound (Ia) at 0.1mg/kg+Metformin at 150 mg/kg has shown increased 45% reduction in serumglucose. The effect on glucose area under curve in OGTT in differenttreatment groups and % change Vs Vehicle Control is depicted in Tableno. 12

TABLE 12 % improvement in AUC AUC Glucose glucose Compounds (mg/dl.min)vs control Vehicle control 56845.5 ± 4313.6 Compound (Ia)—0.1 mg/kg,p.o. 51757.3 ± 3366.3  9.0 ± 5.9 Metfomin—150 mg/kg, p.o. 45255.9 ±4361.1 20.4 ± 7.7 Compound (Ia)—0.1 mg/kg, p.o. + 28176.3 ± 2548.9 50.4± 4.5 Metfomin—150 mg/kg, p.o.

In oral glucose tolerance test, Compound (Ia) at 0.1 mg/kg showed only9% improvement in glucose area under curve and Metformin at 150 mg/kgshowed 20.4% improvement glucose tolerance whereas co-administration ofCompound (Ia) at 0.1 mg/kg+Metformin at 150 mg/kg has shownsynergistically 50.4% improvement in glucose area under curve during theOGTT done after 14 days of treatment, so it indicates thatco-administration of Compound (Ia) and Metformin has shown synergisticactivity in the antidiabetic parameters evaluated.

III) Lipid Lowering Effects of Compound (Ia), Rosuvastatin Calcium andCo-Administration of Compound (Ia)+Rosuvastatin Calcium, in SyrianGolden Hamsters

The in-vivo efficacy was done in high fat high cholesterol fed Syriangolden hamster. It is an excellent model to investigate hepatic assemblyand secretion of lipoprotein since its lipoprotein metabolism closelyresembles that of humans as they have cholesteryl ester transfer protein(CETP) as like human. Feeding high fat high cholesterol diet to Syriangolden hamster causes hypercholesterolemia and hypertriglyceridemiawhich causes an increase in LDL levels and decrease in HDL-C/TC ratio.

In this experiment, 21 Syrian golden hamsters which were kept on highfat-high cholesterol (HF-HC) fructose diet were divided into 3 treatmentgroups and one group was on normal diet control which was NIN (NationalInstitute of Nutrition, NIN diet) control, the treatment given was asper Table no. 1. NIN control and HF-HC fructose diet vehicle controlgroups were treated with vehicle (10% Polyethylene glycol (PEG) 400+90%of 0.5% methyl cellulose) and other three groups with treatmentmentioned in Table no. 13. The treatment was given oral gavages oncedaily for 7 days.

TABLE NO. 13 Dosage Number of Sr. no Treatment Group Level Sex Animals 1NIN control 0 mg/kg M 7 2 HF-HC fructose diet 0 mg/kg M 7 vehiclecontrol 3 Compound (Ia) 3 mg/kg M 7 4 Compound (Ia)   3 mg/kg + M 7 (3mg/kg) + 0.3 mg/kg   Rosuvastatin Calcium (0.3 mg/kg)   

The blood was collected by retro-orbital sinus puncture method underlight ether anesthesia on day 0 (pretreatment) and on day 7 of thetreatment, serum was separated and analyzed for Low density cholesterol(LDL-C) and High density cholesterol (CDL-C) levels using. Cobas C 311clinical chemistry autoanalyser from Roche Diagnostics using commercialdiagnostic kits.

Calculations of the % change in LDL-C levels, HDL-C and LDL-C/HDL-Cratio (Vs HF-HC fructose diet control) were performed using MS Excel.

The serum LDL-C levels in different treatment groups and % change VsHF-HC Fructose Control is depicted in Table no. 14

TABLE 14 % Change LDL-C % vs HF-HC Serum LDL-C (mg/dl) Change FructoseTreatment Day 0 Day 7 vs Day 0 Control NIN control 17.6 ± 1.2 15.2 ± 1.2−13.6 ± 4.5  — HF-FC-Fructose 111.2 ± 6.6  134.8 ± 13.5 24.1 ± 15.2 —control Compound (Ia) 114.8 ± 14.4 123.7 ± 15.5 9.2 ± 7.8 −9.9 ± 6.5 (3mg/kg) Compound (Ia) 115.3 ± 17.5  97.9 ± 13.8 −13.9 ± 3.9  −29.0 ± 3.2 (3 mg/kg) + Rosuvastatin calcium (0.3 mg/kg)

The treatment with Compound (Ia) at 3 mg/kg showed only 9.9% reductionin LDL-C and whereas co-administration of Compound (Ia) (3mg/kg)+Rosuvastatin calcium (0.3 mg/kg) has shown 29% reduction inLDL-C.

The serum HDL-C levels in different treatment groups and % change VsHF-HC Fructose Control is depicted in Table no. 15

TABLE 15 % Change HDL-C vs HF-HC Serum HDL-C (mg/dl) % Change FructoseTreatment Day 0 Day 7 vs Day 0 Control NIN control  64.0 ± 3.1 64.5 ±3.8  1.1 ± 4.9 — HF-FC-Fructose 101.0 ± 5.6 115.2 ± 5.4  15.1 ± 5.4 —control Compound (Ia) 101.8 ± 3.2 111.5 ± 4.3   9.8 ± 4.5 −3.7 ± 3.9 (3mg/kg) Compound (Ia) 101.4 ± 4.5 142.8 ± 10.7 40.0 ± 5.4 22.7 ± 4.7 (3mg/kg) + Rosuvastatin calcium (0.3 mg/kg)

The treatment with Compound (Ia) at 3 mg/kg does not show any increaseHDL-C whereas co-administration of Compound (Ia) (3 mg/kg)+Rosuvastatincalcium (0.3 mg/kg) has shown 22.7% increase in HDL-C.

The serum LDL-C/HDL-C ratio, atherogenic index in different treatmentgroups and % change Vs HF-HC Fructose Control are depicted in Table no.16

TABLE 16 % Change Serum LDL-C/ LDL/HDL vs HF-HC HDL-C Ratio % ChangeFructose Treatment Day 0 Day 7 Vs Day 0 Control NIN control 0.28 ± 0.00.24 ± 0.0 −14.3 ± 2.8  — HF-FC-Fructose 1.11 ± 0.0 1.16 ± 0.1  6.3 ±9.4 — control Compound (Ia) 1.13 ± 0.1 1.09 ± 0.1 −0.3 ± 6.7  −4.9 ± 6.4(3 mg/kg) Compound (Ia) 1.12 ± 0.1 0.67 ± 0.0 −38.1 ± 3.4  −41.0 ± 3.3(3 mg/kg) + Rosuvastatin calcium (0.3 mg/kg)

The treatment with Compound (Ia) at 3 mg/kg showed only 4.9% reductionin LDL-C/HDL-C ratio, atherogenic index whereas co-administration ofCompound (Ia) (3 mg/kg)+Rosuvastatin calcium (0.3 mg/kg) has shownsynergistically 41.0% reduction in LDL-C/HDL-C ratio, atherogenic index.

In clinical trials, Compound of Formula Ia and Atorvastatin combinationshowed significantly better reduction in triglyceride and LDL-C levelsas compared to placebo group.

Further, no side effects associated with PPARs have been seen.

IV) Antidiabetic Activity of Compound (Ia), Co-Administration ofCompound (Ia)+Vildagliptin (Dipeptidyl Peptidase IV Inhibitor) andCo-Administration of Compound (is)+Vildagliptin (Dipeptidyl Peptidase IVInhibitor)+Metformin in db/db Mice Model

The in-vivo efficacy for antidiabetic activity was done female db/dbmice of 10-15 weeks of age. Generally obese and diabetic animals areinsulin resistant and have abnormalities in glucose and lipidmetabolism. The development of diabetes and beta cell dysfunction inthese animal models closely parallels the pathophysiology of the diseasecondition in humans. The db/db mouse exhibits an initial phase ofhyperinsulinemia, hyperphagia and obesity. They progressively developinto insulinopenia with age, a feature commonly observed in last stageof type-2 diabetes, when blood sugar levels are insufficientlycontrolled. Therefore, this genetic model proves to be suitable forpredicting the likely therapeutic benefit of novel euglycemic agents inhuman type-2 diabetes.

In this experiment, 28 female db/db mice of 10-15 week were divided into4 groups and as per their day 0 serum glucose levels per Table no. 1.Vehicle Control group was treated with vehicle (5% Polyethylene glycol(PEG) 400+5% Tween 80+90% of 0.5% Sodium Carboxy methyl cellulose) andother four groups with treatment mentioned in Table no. 17. Thetreatment was given oral gavages once daily for 14 days.

TABLE NO. 17 Dosage Number of Sr. no Treatment Group Level Sex Animals 1Vehicle Control 0 mg/kg Female 7 2 Compound (Ia) 0.1 mg/kg   Female 7 3Compound (Ia) 3 mg/kg Female 7 (0.1 mg/kg) + Vildagliptin (1 mg/kg) 4Compound (Ia) 0.1 mg/kg + Female 7 (0.1 mg/kg) +   1 mg/kg +Vildagliptin 100 mg/kg    (1 mg/kg) + Metformin (100 mg/kg) 

The blood was collected by retro-orbital sinus puncture method underlight ether anesthesia on day 0 (pretreatment) and on day 12 (one hourafter dose administration) for non-fasting glucose analysis and oralglucose tolerance test (OGTT) was performed on day 14 of the treatmentin fasting condition. After blood collection serum was separated andanalyzed glucose analysis using Spectra max and commercial diagnostickits. The OGTT procedure followed is as given below—

Oral Glucose Tolerance Test (OGTT):

One day before OGTT all animals were kept on overnight fasting and onthe day of OGTT, day 14 of the treatment each animal was dosed with asingle dose of vehicle/test compounds administered orally and 60-minpost dosing blood was collected (0 min) and glucose load (2 gm/kg/10 ml)administered per orally. Blood was then collected at time pointscorresponding to 30, 60 and 120 min after glucose load administration.Serum was separated and analyzed for glucose using Spectra max andcommercial diagnostic kits.

The calculation for antidiabetic activity i.e. effect on serum glucoseand improvement in glucose Area Under Curve (AUC) was calculated usingthe excel sheet and Graph Pad Prism software. The day 0 and day 12glucose values of vehicle control and treatment were considered forcalculating the effect on serum glucose. The effect on glucose AUC wascalculated using the OGTT glucose levels at different time points andthe improvement in glucose AUC were the reduction in glucose AUC afterthe treatment during OGTT.

The effect on serum glucose levels in different treatment groups and %change Vs Vehicle Control is depicted in Table no. 18

TABLE 18 % Change Serum Glucose (mg/dl) Day 12 % Change Compound Day 0Day 12 vs Day 0 Vs control Vehicle Control 372.7 ± 31.8 318.9 ± 30.8−11.9 ± 3.8 Compound Ia 375.4 ± 26.8 209.0 ± 18.1 −43.8 ± 3.9 −34.3 ±4.5 (0.1 mg/kg) Compound Ia 389.1 ± 28.0 195.2 ± 12.6 −49.0 ± 3.8 −40.4± 4.4 (0.1 mg/kg, p.o) + Vildagliptin (1 mg/kg, p.o) Compound Ia 365.5 ±35.2 146.8 ± 7.9  −57.2 ± 5.3 −49.9 ± 6.2 (0.1 mg/kg, p.o) +Vildagliptin (1 mg/kg, p.o) + Metformin (100 mg/kg)

The treatment with Compound (Ia) at 0.1 mg/kg showed 34.3% reduction inserum glucose and co-administration of Compound (Ia) at 0.1mg/kg+Vildagliptin (dipeptidyl peptidase IV inhibitor) at 1 mg/kg hasshown 40% reduction in serum glucose whereas co-administration ofCompound (Ia) at 0.1 mg/kg+Vildagliptin (dipeptidyl peptidase IVinhibitor) at 1 mg/kg+Metfomin at 100 mg/kg showed 49.9% reduction inserum glucose.

The effect on glucose area under curve in OGTT in different treatmentgroups and % change Vs Vehicle Control is depicted in Table no. 19

TABLE 19 % improvement AUC Glucose in AUC glucose Compounds (mg/kg)(mg/dl.min) vs control Vehicle Control 62132.2 ± 3749.0 Compound Ia (0.1mg/kg) 69613.3 ± 2547.9 −12.0 ± 4.1  Compound Ia (0.1 mg/kg, p.o) +42351.0 ± 3087.4 31.8 ± 5.0 Vildagliptin (1 mg/kg, p.o) Compound Ia (0.1mg/kg, p.o) + 23963.3 ± 2048.7 61.4 ± 3.3 Vildagliptin (1 mg/kg, p.o) +Metformin (100 mg/kg)

In oral glucose tolerance test, Compound (Ia) at 0.1 mg/kg showed doesno show any improvement in glucose area under curve andco-administration of Compound (Ia) at 0.1 mg/kg+Vildagliptin at 1 mg/kghas shown synergistically 31.8% improvement in glucose area under curve(AUC) during the OGTT done after 14 days of treatment, whereas andco-administration of Compound (Ia) at 0.1 mg/kg+Vildagliptin at 1mg/kg+Metfromin at 100 mg/kg has shown synergistically 61.4% improvementin glucose AUC during the OGTT, so it indicates that co-administrationof Compound (Ia) and Vildagliptin and metfomim has shown synergisticactivity antidiabetic parameters evaluated i.e. serum glucose and oralglucose tolerance test.

V) Lipid Lowering Effects of Compound (La), and Co-Administration ofCompound (Ia)+Atorvastatin, in Syrian Golden Hamsters

The in-vivo efficacy was done in high fat high cholesterol fed Syriangolden hamster. It is an excellent model to investigate hepatic assemblyand secretion of lipoprotein since its lipoprotein metabolism closelyresembles that of humans as they have cholesteryl ester transfer protein(CETP) as like human. Feeding high fat high cholesterol diet to Syriangolden hamster causes hypercholesterolemia and hypertriglyceridemiawhich causes an increase in LDL levels and decrease in HDL-C/TC ratio.

In this experiment, 18 Syrian golden hamsters which were kept on highfat-high cholesterol (HF-HC) fructose diet were divided into 3 treatmentgroups and one group was on normal diet control which was NIN (NationalInstitute of Nutrition, NIN diet) control, the treatment given was asper Table no. 1. NIN control and HF-HC fructose diet vehicle controlgroups were treated with vehicle (5% Polyethylene glycol (PEG) 400+5%Tween 80+90% of 0.5% Sodium Carboxy methyl cellulose) and other threegroups with treatment mentioned in Table no. 20. The treatment was givenoral gavages once daily for 13 days.

TABLE NO. 20 Dosage Number of Sr. no Treatment Group Level Sex Animals 1NIN. control 0 mg/kg M 6 2 HF-HC fructose diet 0 mg/kg M 6 vehiclecontrol 3 Compound (Ia) 3 mg/kg M 6 4 Compound (Ia)   3 mg/kg + M 6   (3mg/kg) + 0.5 mg/kg   Atorvastatin (0.5 mg/kg)

The blood was collected by retro-orbital sinus puncture method underlight ether anesthesia on day 0 (pretreatment) and on day 13 of thetreatment, serum was separated and analyzed for Low density cholesterol(LDL-C) and High density cholesterol (CDL-C) and triglyceride levelsusing Cobas C 311 clinical chemistry autoanalyser from Roche Diagnosticsusing commercial diagnostic kits.

Calculations of the % change in LDL-C levels, triglyceride andLDL-C/HDL-C ratio (Vs HF-HC fructose diet control) were performed usingMS Excel.

The serum LDL-C levels in different treatment groups and % change VsHF-HC Fructose Control is depicted in Table no. 21

TABLE 21 % Change Serum LDL-C vs HF-HC LDL-C (mg/dl) % Change FructoseTreatment Day 0 Day 13 vs Day 0 Control NIN control 11.3 ± 0.6 10.7 ±0.7  −5.5 ± 3.9 HF-HC-Fructose 74.1 ± 4.0 87.1 ± 10.0 16.1 ± 8.4 controlCompound Ia 75.1 ± 6.9 73.1 ± 4.3   0.1 ± 7.6 −14.8 ± 6.5 (3 mg/kg)Compound Ia 79.3 ± 5.4 55.6 ± 5.1  −28.0 ± 9.1  −38.7 ± 7.7 (3 mg/kg) +Atorvastatin 0.5 mg/kg

The treatment with Compound (Ia) at 3 mg/kg showed only 14.8% reductionin LDL-C whereas co-administration of Compound (Ia) (3mg/kg)+Atorvastatin (0.5 mg/kg) has shown 38.7% reduction in LDL-C.

The effect on triglyceride in different treatment groups and % change VsHF-HC Fructose Control is depicted in Table no. 22

TABLE 22 % Change Serum TG % vs HF-HC triglyceride (mg/dl) ChangeFructose Treatment Day 0 Day 13 vs Day 0 Control NIN control 117.7 ±6.1  153.5 ± 15.4 30.2 ± 9.6 HF-HC-Fructose 446.4 ± 32.0 461.7 ± 48.2 6.2 ± 13.0 control Compound Ia 418.4 ± 35.2 149.5 ± 25.8 −64.0 ± 5.1 −65.2 ± 4.9 (3 mg/kg) Compound Ia 434.8 ± 35.7 79.7 ± 9.2 −81.0 ± 2.7 −81.6 ± 2.6 (3 mg/kg) + Atorvastatin 0.5 mg/kg

The treatment with Compound (Ia) at 3 mg/kg showed 65.2% reduction intriglyceride whereas co-administration of Compound (Ia) (3mg/kg)+Atorvastatin (0.5 mg/kg) has shown 81.6% reduction intriglyceride The serum LDL-C/HDL-C ratio, atherogenic index in differenttreatment groups and % change Vs HF-HC Fructose Control are depicted inTable no. 23

TABLE 23 % Change vs LDL/HDL HF-HC RATIO Change Fructose Treatment Day 0Day 13 vs Day 0 Control NIN control 0.19 ± 0.0 0.15 ± 0.0 −21.9 ± 5.4 HF-HC-Fructose 0.78 ± 0.1 0.77 ± 0.1 −2.4 ± 4.8 control Compound Ia 0.79± 0.1 0.82 ± 0.0  6.4 ± 7.4  8.5 ± 7.6 (3 mg/kg) Compound Ia 0.88 ± 0.10.61 ± 0.1 −26.6 ± 11.1 −25.1 ± 11.4 (3 mg/kg) + Atorvastatin 0.5 mg/kgThe treatment with Compound (Ia) at 3 mg/kg does not showed anyreduction in LDL-C/HDL-C ratio, atherogenic index whereasco-administration of Compound (Ia) (3 mg/kg)+Atorvastatin (0.5 mg/kg)has shown synergistically 25.1% reduction in LDL-C/HDL-C ratio,atherogenic index.

Therefore, the compounds of formula (Ia) shows synergistic effect withseveral other therapeutic agents as described in the specification.

VI) Antidiabetic Activity of Metformin+Insulin and Co-Administration ofCompound (Ia)+Metformin+Insulin in db/db Mice Model.

The in-vivo efficacy for antidiabetic activity was done male db/db miceof 12-15 weeks of age. Generally obese and diabetic animals are insulinresistant and have abnormalities in glucose and lipid metabolism. Thedevelopment of diabetes and beta cell dysfunction in these animal modelsclosely parallels the pathophysiology of the disease condition inhumans. The db/db mouse exhibits an initial phase of hyperinsulinemia,hyperphagia and obesity. They progressively develop into insulinopeniawith age, a feature commonly observed in last stage of type-2 diabetes,when blood sugar levels are insufficiently controlled. Therefore, thisgenetic model proves to be suitable for predicting the likelytherapeutic benefit of novel euglycemic agents in human type-2 diabetes.

In this experiment, 21 male db/db mice of 12-15 week were divided into 3groups and as per their day 0 serum glucose levels. Vehicle Controlgroup was treated with vehicle (5% Polyethylene glycol (PEG) 400+5%Tween 80+90% of 0.5% Sodium Carboxy methyl cellulose) and other 2 groupswith treatment mentioned in Table no. 24. Compound (Ia) and metforminwas administered per orally and insulin was injected subcutaneously asper dose given in table 24, once in day for 14 days.

TABLE NO. 24 Dosage Number of Sr. no Treatment Group Level Sex Animals 1Vehicle Control   0 mg/kg male 7 2 Metformin 100 mg/kg, male 7 (100mg/kg, p.o) + per oral + Insulin 0.1 U/mouse, s.c. (0.1 U/Mouse s.c) 3Metformin 100 mg/kg, male 7 (100 mg/kg, p.o) + per oral + Insulin   0.1U/mouse, s.c. +   (0.1 U/Mouse s.c) + 0.1 mg/kg Compound (Ia) per oral  (0.1 mg/kg, p.o.) +The blood was collected by retro-orbital sinus puncture method underlight ether anesthesia on day 0 (pretreatment) and on day 12 (one hourafter dose administration) for non-fasting glucose analysis and oralglucose tolerance test (OGTT) was performed on day 14 of the treatmentin fasting condition. After blood collection serum was separated andanalyzed glucose analysis using Spectra max and commercial diagnostickits. The OGTT procedure followed is as given below—

Oral Glucose Tolerance Test (OGTT):

One day before OGTT all animals were kept on overnight fasting and onthe day of OGTT, day 14 of the treatment each animal was dosed with asingle dose of vehicle/test compounds administered orally and 60-minpost dosing blood was collected (0 min) and glucose load (2 gm/kg/10 ml)administered per orally. Blood was then collected at time pointscorresponding to 30, 60 and 120 min after glucose load administration.Serum was separated and analyzed for glucose using Spectra max andcommercial diagnostic kits.

The calculation for antidiabetic activity i.e. effect on serum glucoseand improvement in glucose Area Under Curve (AUC) was calculated usingthe excel sheet and Graph Pad Prism software. The day 0 and day 12glucose values of vehicle control and treatment were considered forcalculating the effect on serum glucose. The effect on glucose AUC wascalculated using the OGTT glucose levels at different time points andthe improvement in glucose AUC were the reduction in glucose AUC afterthe treatment during OGTT.

The effect on serum glucose levels in different treatment groups and %change Vs Vehicle Control is depicted in Table no. 25

TABLE 25 Serum % Change Glucose (mg/dl) Day 12 % Change Compound Day 0Day 12 vs Day 0 Vs control Vehicle (p.o) 523.8 ± 30.2 527.5 ± 36.1  1.3± 6.4 Metformin 522.4 ± 32.4 212.5 ± 24.0 −58.1 ± 5.3 −58.4 ± 5.3 (100mg/kg, p.o) + Insulin (0.1 U/Mouse s.c) Metformin 532.1 ± 35.8 126.0 ±18.3 −75.4 ± 4.1 −75.6 ± 4.0 (100 mg/kg, p.o) + Insulin (0.1 U/mouse,s.c) + Compound Ia (0.1 mg/kg, p.o)

The treatment with Metformin+insulin showed 58.4% reduction in serumglucose and co-administration of Compound (Ia) at 0.1 mg/kg along withMetformin+insulin has shown 75.6% reduction in serum glucose. Incombination Compound (Ia) has caused significant increase inantidiabetic activity of Metformin+insulin.

The effect on glucose area under curve in OGTT in different treatmentgroups and % change vs Vehicle Control is depicted in Table no. 26

TABLE 26 Day 14 % improvement AUC Glucose in AUC Compounds (mg/kg)(mg/dl.min) glucose vs control Vehicle (p.o) 69455.4 ± 3670.6 Metformin(100 mg/kg, p.o) + 30743.6 ± 3689.9 55.7 ± 5.3 Insulin (0.1 U/Mouse s.c)Metformin (100 mg/kg, p.o) + 27675.3 ± 1984.3 60.2 ± 2.9 Insulin (0.1U/mouse, s.c) + Compound Ia (0.1 mg/kg, p.o)

In oral glucose tolerance test, treatment with Metformin+insulin showed55.7% improvement in glucose area under curve whereas co-administrationof Compound (Ia) at 0.1 mg/kg along with Metformin+insulin has shown60.2% improvement in glucose area under curve (AUC) during the OGTT doneafter 14 days of treatment. This indicates that co-administration ofCompound (Ia) and Metformin+insulin has increased the antidiabeticactivity in parameters evaluated i.e. serum glucose and oral glucosetolerance test.

VII) Antidiabetic Activity of Compound (in) and Co-Administration ofCompound (Ia)+Exenatide (GLP-1 Receptor Agonist) in db/db Mice Model.

VIII) Antidlabetic Activity of Compound (Ia), Co-Administration ofCompound (Ia)+Glimepiride (Sulfonylurea) in db/db Mice Model.

The in-vivo efficacy for antidiabetic activity was done in male andfemale db/db mice of 12-16 weeks of age. Generally obese and diabeticanimals are insulin resistant and have abnormalities in glucose andlipid metabolism. The development of diabetes and beta cell dysfunctionin these animal models closely parallels the pathophysiology of thedisease condition in humans. The db/db mouse exhibits an initial phaseof hyperinsulinemia, hyperphagia and obesity. They progressively developinto insulinopenia with age, a feature commonly observed in last stageof type-2 diabetes, when blood sugar levels are insufficientlycontrolled. Therefore, this genetic model proves to be suitable forpredicting the likely therapeutic benefit of novel euglycemic agents inhuman type-2 diabetes.

In this experiment, 49 db/db mice of 12-16 week were divided into 7groups and as per their day 0 serum glucose levels. Exenatide wasadministered intraperitoneally (i.p.) after diluting in phosphate buffersaline (PBS) and Compound Ia and glimepiride was administered per orallyso there was two vehicle control group one was intraperitoneally (i.p.)PBS for comparing with exenatide i.p. group and another vehicle controlgroup was treated with vehicle used for oral administration (5%Polyethylene glycol (PEG) 400+5% Tween 80+90% of 0.5% Sodium Carboxymethyl cellulose) of Compound Ia and glimepiride. Detail treatments androutes of administrations were mentioned in Table no. 27. The treatmentwas given once daily for 7 days.

TABLE NO. 27 Dosage Level and route Sr. no Treatment Group ofadministration Number of Animals 1 Vehicle Control for I.P, 0 mg/kg 3female + 4 Male = 7 ((PBS-10 ml/kg, i.p) 2 Exenatide (0.1 μg/kg, i.p)0.1 μg/kg, i.p 3 female + 4 Male = 7 3 Vehicle Control for per oral 0mg/kg, p.o. 3 female + 4 Male = 7 (10 ml/kg, p.o) 4 Compound Ia (0.1mg/kg. p.o.)   0.1 mg/kg. p.o. 4 female + 3 Male = 7 5 Glimepiride (2mg/kg, p.o.)   2. mg/kg, p.o 3 female + 4 Male = 7 6 Compound Ia (0.1mg/kg, p.o) +  0.1 mg/kg. p.o. + 3 female + 4 Male = 7 Exenatide (0.1μg/kg, i.p) 0.1 μg/kg, i.p 7 Compound Ia (0.1 mg/kg, p.o) +  0.1 mg/kg.p.o.+ 3 female + 4 Male = 7 Glimepiride (2 mg/kg, p.o)  2 mg/kg, p.o.

The blood was collected by retro-orbital sinus puncture method underlight ether anesthesia on day 0 (pretreatment) and on day 6 (one hourafter dose administration) for non-fasting glucose analysis and oralglucose tolerance test (OGTT) was performed on day 7 of the treatment infasting condition. After blood collection serum was separated andanalyzed glucose analysis using Spectra max and commercial diagnostickits. The OGTT procedure followed is as given below—

Oral Glucose Tolerance Test (OGTT):

One day before OGTT all animals were kept on overnight fasting and onthe day of OGTT, day 7 of the treatment each animal was dosed with asingle dose of vehicle/test compounds administered orally and 60-minpost dosing blood was collected (0 min) and glucose load (2 gm/kg/10 ml)administered per orally. Blood was then collected at time pointscorresponding to 30, 60 and 120 min after glucose load administration.Serum was separated and analyzed for glucose using Spectra max andcommercial diagnostic kits.

The calculation for antidiabetic activity i.e. effect on serum glucoseand improvement in glucose Area Under Curve (AUC) was calculated usingthe excel sheet and Graph Pad Prism software. The day-0 and day-6glucose values of vehicle control and treatment were considered forcalculating the effect on serum glucose. The effect on glucose AUC wascalculated using the OGTT glucose levels at different time points andthe improvement in glucose AUC were the reduction in glucose AUC afterthe treatment during OGTT.

The effect on serum glucose levels after treatment with Exenatide andits combination on day 6 in various groups and % change Vs VehicleControl is depicted in Table no. 28

TABLE 28 % Change Serum Glucose (mg/dl) Day 6 vs % Change Compound Day 0Day 6 Day 0 Vs control Vehicle for I.p. 415.7 ± 24.7 457.8 ± 40.5  13.7± 14.5 (PBS-10 ml/kg, i.p) Exenatide 408.4 ± 23.2 317.3 ± 25.0 −22.2 ±4.6 −29.3 ± 4.2 (0.1 μg/kg, i.p) Vehicle for 405.1 ± 23.1 466.6 ± 30.9 15.0 ± 2.8 per oral (10 ml/kg, p.o) Compound Ia 420.6 ± 25.1 363.4 ±39.0 −13.4 ± 8.1 −24.8 ± 7.0 (0.1 mg/kg, p.o) Compound Ia 402.5 ± 22.6224.0 ± 28.9 −43.0 ± 6.1 −50.5 ± 5.3 (0.1 mg/kg, p.o) + Exenatide (0.1μg/kg, i.p)

The treatment with Compound Ia (0.1 mg/kg, p.o.) and Exenatide (0.1μg/kg, i.p.) showed 24.8 and 29.3% reduction in serum glucose whereasco-administration of Compound (Ia) at 0.1 mg/kg+Exenatide (GLP-1receptor agonist) at 0.1 μg/kg has shown 50.5% reduction in serumglucose.

The effect on glucose area under curve in OGTT after treatment withExenatide and its combination and % change Vs Vehicle Control isdepicted in Table no. 29

TABLE 29 % improvement in AUC AUC Glucose glucose vs Compounds (mg/kg)(mg/dl.min) control Vehicle for I.p. 50202.8 ± 3020.2 (PBS-10 ml/kg,i.p) Exenatide (0.1 μg/kg, i.p) 40025.6 ± 4392.0 20.3 ± 8.7 Vehicle forper oral 60917.9 ± 3892.3 (10 ml/kg, p.o) Compound Ia 46452.6 ± 4298.123.7 ± 7.1 (0.1 mg/kg, p.o) Compound Ia 33143.7 ± 4591.8 45.6 ± 7.5 (0.1mg/kg, p.o) + Exenatide (0.1 μg/kg, i.p)

In oral glucose tolerance test, treatment with Compound Ia (0.1 mg/kg,p.o.) and Exenatide (0.1 μg/kg, i.p.) showed 23.7 and 20.3% improvementin glucose area under curve (AUC) whereas co-administration of Compound(Ia) at 0.1 mg/kg+Exenatide (GLP-1 receptor agonist) at 0.1 μg/kg hasshown 45.6% improvement in glucose AUC during the OGTT.

This indicates that co-administration of Compound (Ia) and Exenatide(GLP-1 receptor agonist) has shown synergistic activity in antidiabeticparameters evaluated i.e. serum glucose and oral glucose tolerance test.

The effect on serum glucose levels after treatment with Glimepiride andits combination on day 6 in various groups and % change Vs VehicleControl is depicted in Table no. 30

TABLE 30 % Change Serum Glucose (mg/dl) Day 6 vs % Change Compound Day 0Day 6 Day 0 Vs control Vehicle Control 405.1 ± 23.1 466.6 ± 30.9  15.0 ±2.8 (10 ml/kg, p.o) Compound Ia 420.6 ± 25.1 363.4 ± 39.0 −13.4 ± 8.1−24.8 ± 7.0  (0.1 mg/kg, p.o) Glimepiride 402.0 ± 25.8 506.7 ± 34.8 26.9 ± 7.3 10.1 ± 6.4 (2 mg/kg, p.o) Compound Ia 357.3 ± 33.9 247.0 ±40.0  −32.6 ± 12.6 −41.4 ± 10.9 (0.1 mg/kg, p.o) + Glimepiride (2 mg/kg,po)

The treatment with Compound Ia (0.1 mg/kg, p.o.) showed 24.8% reductionin serum glucose but glimepiride (2 mg/kg, p.o.) showed 10.1% increasein serum glucose as compared to vehicle control. Whereasco-administration of Compound (Ia) at 0.1 mg/kg+glimepiride(sulfonylurea) at 2 mg/kg, p.o. has shown synergistically 41.4%reduction in serum glucose.

The effect on glucose area under curve (AUC) in OGTT after treatmentwith Glimepiride and its combination in different treatment groups and %change Vs Vehicle Control is depicted in Table no. 31

TABLE 31 % improvement AUC Glucose in AUC glucose Compounds (mg/kg)(mg/dl.min) vs control Vehicle for per oral (10 ml/kg, p.o) 60917.9 ±3892.3 Compound Ia (0.1 mg/kg, p.o) 46452.6 ± 4298.1 23.7 ± 7.1Glimepiride (2 mg/kg, p.o) 62529.0 ± 3008.2 −2.6 ± 4.9 Compound Ia (0.1mg/kg, p.o) + 45079.3 ± 5367.3 26.0 ± 8.8 Glimepiride (2 mg/kg, po)

In oral glucose tolerance test, Compound (Ia) at 0.1 mg/kg showed 23.7%improvement in glucose area under curve and glimepiride (2 mg/kg, p.o.)does not any improvement in glucose AUC. Whereas co-administration ofCompound (Ia) at 0.1 mg/kg+glimepiride (2 mg/kg, p.o.) has shownsynergistically 26% improvement in glucose area under curve (AUC) duringthe OGTT done after 7 days of treatment.

This indicates that co-administration of Compound (Ia) and glimepiride(sulfonylurea) has shown synergistic activity in antidiabetic parametersevaluated i.e. serum glucose and oral glucose tolerance test.

1. Synergistic composition comprising a compound of formula (Ia)

wherein ‘M⁺’ represents Calcium, Magnesium, Sodium, Potassium, Zinc andLithium and one or more therapeutic agent selected from one or more DPPIV inhibitors, one or more biguanide antihyperglycaemic agents, one ormore statins, one or more thiazolidinediones, one or more sulfonylureas,one or more SGLT2 inhibitors, one or more insulin sensitizers or one ormore GLP-1 agonists.
 2. The synergistic composition as claimed in claim1, wherein the one or more DPP IV inhibitors are selected fromSitagliptin, Vildagliptin, Saxagliptin, Alogliptin and Linagliptin. 3.The synergistic composition as claimed in claim 1, wherein the one ormore statins are selected from Lovastatin, Pravastatin, Fluvastatin,Simvastatin, Atorvastatin, Rosuvastatin and Pitavastatin.
 4. Thesynergistic composition as claimed in claim 1, wherein the one or morebiguanide antihyperglycaemic agents are selected from Metformin,Buformin or Phenformin.
 5. The synergistic composition as claimed inclaim 1, wherein the one or more thiazolidinediones are selected fromPioglitazone and Rosiglitazone.
 6. The synergistic composition asclaimed in claim 1, wherein the one or more sulphonylureas are selectedfrom glibenclamide, glipizide, gliclazide, glimepiride, tolazamide,tolbutamide, acetohexamide, carbutamide, chlorpropamide, glibomuride,gliquidone, glisentide, glisolamide, glisoxepide, glyclopyamide,glycylamide and glipentide.
 7. The synergistic composition as claimed inclaim 1, wherein the one or more SGLT-2 inhibitors are selected fromDapagliflozin, Canagliflozin, Empagliflozin, Ertugliflozin andIpragliflozin.
 8. The synergistic composition as claimed in claim 1,wherein the one or more GLP-1 agonists are selected from Exenatide,Liraglutide and Dulaglutide.
 9. (canceled)
 10. A method for thetreatment of dyslipidemia, hypertriglyceridemia or diabetes mellituswhich comprises administering an effective, non-toxic andpharmaceutically acceptable amount of a composition according to claim1, mammal in need thereof.
 11. A composition according to claim 1,further comprising another suitable therapeutic agent for the treatmentobesity, diabetes mellitus, especially Type 2 diabetes and conditionsassociated with diabetes mellitus.
 12. The composition as claimed inclaim 11, wherein the another suitable therapeutic agent is selectedfrom an antidiabetic agent, an alpha glucosidase inhibitor, a biguanide,an insulin secretagogue or an insulin sensitizer and sulphonylureas. 13.The composition according to claim 1, further comprising one or morepharmaceutically acceptable excipients.
 14. (canceled)
 15. Thepharmaceutical composition as claimed in claim 13 in the form of atablets, capsules, powders, granules, lozenge, suppositories,reconstitutable powder, or liquid preparations.
 16. (canceled) 17.(canceled)
 18. The composition as claimed in claim 13, wherein thepharmaceutically acceptable excipient is selected from suitable bindingagents, fillers, lubricants, glidants, disintegrants and wetting agents.19. (canceled)
 20. A compound of Formula (I)

wherein R is selected from —SCH₃ or —OCH₃, and M⁺ is selected from thegroup consisting of Calcium, Sodium, Potassium, Zinc, Lithium,L-Arginine, Tromethamine, L-Lysine, Meglumine, Benethamine, Piperazine,Benzylamine, Dibenzylamine, Dicyclohexylamine, Diethylamine,Diphenylamine, α-naphthylamine, O-phenylenediamine, 1,3-Diaminopropane,(S)-α-naphthylethylamine, (S)-3-methoxyphenylethylamine,(S)-4-methoxyphenylethylamine, (S)-4-chlorophenylethylamine,(S)-4-methylphenylethylamine, Cinchonine, Cinchonidine, (−)-Quinine,Benzathine, Ethanolamine, Diethanol amine, Triethanolamine, imidazole,Diethylamine, Ethylenediamine, Choline, Epolamine, Morpholine4-(2-hydroxyethyl), N-N-diethylethanolamine, Deanol, Hydrabamine,Betaine, Ammonia, Adamantanamine, L-Adamantanmethylamine Tritylamine,Glucamine N-methyl, and Pyrrolidine.
 21. The compound as claimed inclaim 20, in crystalline, amorphous or partially crystalline form. 22.The compound as claimed in claim 20, in solvate or hydrate form.
 23. Thecompound of formula (I) as claimed in claim 20 selected from i. Calciumsalt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; ii. Sodium salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; iii. Potassium salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (iv) Zinc salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (v) Lithium salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (vi) Piperazine salt of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (vii) Tromethamine salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (viii) L-Lysine salt of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (ix) Meglumine salt of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (x) Benethamine salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (xi) Benzylamine salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (xii) Dibenzylamine salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; (xiii) Arginine salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid; and (xiv) Imidazole salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid.
 24. The benethamine salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid as claimed in claim 23 having an XRD) pattern as depicted in FIG.10 and having a 2 theta values of ±0.2 degree at about 10.0, 10.3, 14.5,15.1, 15.7, 16.7, 17.4, 17.9, 18.6, 19.2, 19.8, 21.3, 23.2 and 25.7. 25.The benethamine salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid as claimed in claim 23 having an XRD pattern as depicted in FIG. 11and having a 2 theta values at about 14.8, 16.8, 17.5, 18.3, 19.3, 20.8,22.6 and 24.2±0.2 degree.
 26. The dibenzylamine salt of(S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid as claimed in claim 23 having an XRD pattern as depicted in FIG. 12and having a 2 theta values of about 8.72, 16.8, 18.5, 19.1, 19.6, 20.6,21.6, 22.5 and 24.5±0.2 degree.
 27. The imidazole salt of (S)2-Ethoxy-3-(4-{2-[2-methyl-5-(4-methylthiophenyl)-pyrrol-1-yl]-ethoxy}-phenyl)-propionicacid as claimed in claim 23 having an XRD pattern as depicted in FIG. 14and having a 2 theta values of about 9.40, 14.7, 15.6, 17.3, 21.0, 21.5,22.5 and 26.2±0.2 degree.